Shuttling SR proteins: more than splicing factors

REVIEW ARTICLE

Shuttling SR proteins: more than splicing factorsLaure Twyffels1, Cyril Gueydan1 and Veronique Kruys1,2

1 Laboratoire de Biologie Moleculaire du Gene, Faculte des Sciences, Universite Libre de Bruxelles, Gosselies, Belgium

2 Center of Microscopy and Molecular Imaging, Gosselies, Belgium

Keywords

cytoplasmic; NMD; nucleocytoplasmic;

RNA; shuttling; splicing; SR proteins; SRSF;

sumoylation; translation

Correspondence

V. Kruys, Laboratoire de Biologie

Moleculaire du Gene, Institut de Biologie et

de Medecine Moleculaires, Universite Libre

de Bruxelles, 12 rue des Profs Jeener et

Brachet, 6041 Gosselies, Belgium

Fax: +322 650 9800

Tel: +322 650 9804

E-mail: [email protected]

(Received 20 May 2011, revised 21 June

2011, accepted 25 July 2011)

doi:10.1111/j.1742-4658.2011.08274.x

Serine–arginine (SR) proteins commonly designate a family of eukaryotic

RNA binding proteins containing a protein domain composed of several

repeats of the arginine–serine dipeptide, termed the arginine–serine (RS)

domain. This protein family is involved in essential nuclear processes such

as constitutive and alternative splicing of mRNA precursors. Besides par-

ticipating in crucial activities in the nuclear compartment, several SR pro-

teins are able to shuttle between the nucleus and the cytoplasm and to

exert regulatory functions in the latter compartment. This review aims at

discussing the properties of shuttling SR proteins with particular emphasis

on their nucleo-cytoplasmic traffic and their cytoplasmic functions. Indeed,

recent findings have unravelled the complex regulation of SR protein

nucleo-cytoplasmic distribution and the diversity of cytoplasmic mecha-

nisms in which these proteins are involved.

SR proteins: definition and generalfeatures

Serine–arginine (SR) proteins were first described as a

family of co-purifying proteins capable of restoring

splicing in S100 splicing-deficient extracts. These

factors are structurally highly related as they are com-

posed of a carboxy-terminal domain enriched with the

arginine–serine (RS) dipeptide, preceded by at least

one RNA binding domain of the RNA recognition

motif (RRM) type [1]. These proteins play an essential

role in constitutive and alternative splicing of mRNA

precursors [2]. In the last 10 years, however, other

functions have been unravelled for these proteins.

Indeed, SR protein prototypes such as SRSF1

(ASF ⁄SF2) and SRSF2 (SC35) were reported to stimu-

late transcriptional elongation (for a review see [3]).

Certain SR proteins participate in mRNA translation

(see [4] and below). Finally, a recent study indicates

that SRSF1 promotes microRNA processing by facili-

tating Drosha-mediated cleavage [5]. Altogether, these

findings highlight the broader roles of SR proteins in

gene expression.

Genome-wide searches for proteins containing RS

domains revealed several other ‘non-classical’ SR

proteins, termed SR-like proteins because of differ-

ences in the structure of the RS domain and ⁄or lack of

a prototypical RRM. While a large subset of SR-like

proteins is involved in pre-mRNA processing, several

members of this protein family are associated with

other cellular functions [4,6].

‘Classical’ SR proteins exist in plants [7], metazoans

[1] and in some unicellular eukaryotes, such as the

fission yeast Schizosaccharomyces pombe [8,9]. How-

ever, they are not present in all eukaryotes and are

Abbreviations

ARE, AU-rich element; NMD, nonsense-mediated decay; PIAS1, protein inhibitor of activated STAT-1; RRM, RNA recognition motif;

RS, arginine–serine; SG, stress granule; SR, serine–arginine; UTR, untranslated region.

3246 FEBS Journal 278 (2011) 3246–3255 ª 2011 The Authors Journal compilation ª 2011 FEBS

apparently missing from the budding yeast Saccharo-

myces cerevisiae, which lacks alternative splicing. In

this species, splicing is activated by SR-like proteins

such as Npl3p [10].

To clarify the boundaries of the SR protein family,

a new definition based solely on sequence criteria has

recently been proposed. According to this new defini-

tion, SR proteins contain one or two conserved RNA

binding domains followed by a carboxy-terminal RS

domain of at least 50 amino acids with > 40% RS

content. In human, this strict definition yields a family

of 12 SR proteins [11] (see Fig. 1).

An important question is whether SR proteins have

unique or redundant functions. The intrinsic capacity

of SR proteins to activate splicing was first revealed

by complementation assays with cytoplasmic S100

splicing-deficient extract that contains all the splicing

machinery except SR proteins [1]. However, several

differences distinguish these proteins as to their ability

to promote splicing both in vitro and in vivo, support-

ing unique functions in splicing processes [12–15]. In

Caenorhabditis elegans, gene inactivation of SR

protein homologues by dsRNA interference reveals

that partial functional redundancy ensures viability

upon inactivation of single SR proteins, except in the

case of ASF ⁄SF2 whose expression is essential for

nematode development [16]. The function of SR

proteins has also been studied in mouse model sys-

tems. All SR-null mice for SRSF1, SRSF2 or SRSF3

(SRp20) show an early embryonic phenotype indicat-

ing that SR proteins are not redundant. However, the

essential functions exerted by each SR protein appear

to be tissue and ⁄or developmental stage specific (see

[17] for a review).

SR proteins ensure the coupling of splicing to tran-

scription as they are recruited together with U1

snRNP by RNA polymerase II and co-transcription-

ally deposited on the exon ⁄ 5¢ splice sites of nascent

transcripts [15]. Besides promoting the splicing process,

the co-transcriptional deposition of SR proteins on

nascent pre-mRNA transcripts has been shown to pre-

vent the formation of R loops due to hybridization of

the neosynthesized RNA to the complementary strand

of the DNA template and thereby to contribute to

genome stability by avoiding DNA double-stranded

breaks [18].

Interestingly, SRSF1 was recently reported as a

modulator of protein sumoylation. SRSF1 associates

with Ubc9 and enhances sumoylation of specific

substrates. In addition, SRSF1 interacts with PIAS1

(protein inhibitor of activated STAT-1), regulating

PIAS1-induced overall protein sumoylation. SRSF1

plays a role in heat-shock-induced sumoylation and

promotes SUMO conjugation to RNA-processing fac-

tors. This additional role of SRSF1 further expands

the versatility of this multifunctional protein [19] and

raises the interesting possibility that this activity could

be carried out by other SR proteins.

The expression of SR proteins varies widely among

cell types [20] and it has become clear that the abun-

dance of SR proteins greatly conditions the splicing

programme executed within different cell types. This is

clearly exemplified by SRSF1 which is a proto-onco-

gene whose expression is upregulated in various human

RRM 1 RRM 2 RS

RRM RS

RRM RS

RRM 1 RRM 2 RS

RRM 1 RRM 2 RS

RRM 1 RS

ZnRRM RS

RRM

RRM 2

RS

RRM

RRM 2 RS

RS

RRM RS

RRM RS

RRM

Name Synonym UniProt

SRSF1 (ASF/SF2) Q07955

SRSF2 (SC35) Q01130

SRSF3 (SRp20) P84103

SRSF4 (SRp75) Q08170



SRSF7 (9G8) Q16629

SRSF8 (SRp46) Q9BRL6

SRSF9 (SRp30c) Q13242

SRSF10 (SRp38,SRrp40) O75494

SRSF11 (p54) Q05519

SRSF12 (SRrp35) Q8WXF0

Fig. 1. Schematic representation of the 12

human SR proteins as defined by Manley

and Krainer [11]. Zn, zinc finger.

L. Twyffels et al. Shuttling SR proteins

FEBS Journal 278 (2011) 3246–3255 ª 2011 The Authors Journal compilation ª 2011 FEBS 3247

tumours, partly because of gene amplification. Overex-

pression of SRSF1 leads to abnormal accumulation of

alternatively spliced transcripts, including the mRNA

encoding the oncogenic isoform 2 of ribosomal protein

S6 kinase (S6K1) [21]. In contrast, several autoregula-

tory mechanisms mostly acting at the post-transcrip-

tional level maintain homeostatic levels of SR proteins.

A common mechanism relies on the generation of

alternatively spliced non-productive transcripts which

are either retained in the nucleus or targeted to

nonsense-mediated decay (NMD) due to the inclusion

of premature termination codons [22–24]. This splicing

reprogramming leads to a decrease of the translation-

competent mRNA isoform and thereby to the

repression of SR protein expression. Recent reports

indicate that the expression of SR proteins such as

SRSF1 can also be downmodulated at the post-tran-

scriptional level by microRNAs targeting the SRSF1 3¢untranslated region (UTR) [5,25] or by the autoregula-

tory translational repression of SRSF1 of its own

transcript ([23], and see later). Furthermore, the RS

domain of SR proteins is extensively phosphorylated

on serine residues and this post-translational modifica-

tion plays an important role in regulating the subcellu-

lar localization and activities of SR proteins. Several

protein kinase families have been shown to phosphory-

late the RS domain of SR proteins, including the

SRPK family [26,27], the Clk ⁄Sty family of dual

specificity kinases [28], the DNA topoisomerase I [29]

and the mitogen-activated AKT kinase [30,31]. More

recently, arginine methylation was also reported to

regulate SRSF1 nucleo-cytoplasmic distribution ([32]

and later). All these mechanisms contribute to the fine-

tuning of SR protein functions in various cell types

and upon different cellular conditions.

SR protein localization andnucleo-cytoplasmic shuttling

SR proteins are predominantly located in the nucleus

[33–39] and their nuclear import is mediated by trans-

portin-SR, a member of the b-karyopherin family. This

importin specifically contacts the RS domain to convey

them through the nuclear pore [40,41]. Several lines of

Splicing

Genome stability

Translational control

Localization intostress granules

Transcriptional elongation

mRNA export

SRSF1

SRSF1

SRPK

RISC

(A)nm7g

(A)nm7g

SRSF1 (A)nm7g SRSF1 (A)nm7g

(A)nm7g

+ +

SRSF1

Trn-SR1

Trn-SR2

mTOR+

target targetSUMO

SUMO

SumoylationE1/E2/E3

SRSF1

+

Nonsense-mediated decay

mRNP remodelling

PTC (A)nm7gSRSF1

?

PKCI-r

P

PP

SRSF1+ ? + ?

mRNA degradation

– –

pre-miR-7

pri-miR-7

Pri-miRNAprocessingDrosha

?SRSF1

SRSF1

P

SRSF1

SRSF1

(A)n

(A)n

m7g

m7g

(A) n

m7 g

SRSF1

TIAR

?SRSF1

ESE

?

Xpo5NXF1 NXF1

Fig. 2. Roles of SRSF1 in various cellular processes and compartments. ESE, exonic splicing enhancer; NXF1, nuclear export factor 1; P,

phosphate group; RISC, RNA-induced silencing complex; Xpo5, exportin 5.

Shuttling SR proteins L. Twyffels et al.


evidence demonstrate that phosphorylation of the RS

domain by the cytoplasmic SR protein kinase 1

(SRPK1) is a prerequisite to SR protein nuclear

import [41–43].

Heterokaryon assays revealed that at least four SR

proteins (SRSF1, SRSF3, SRSF7 and SRSF10) shuttle

between the nucleus and the cytoplasm at a high

dynamic rate [34,44,45]. SRSF4 and SRFS6 also shuttle

between the two compartments but with slower kinetics

[44]. Earlier reports indicate that the RS domain is

important for nucleo-cytoplasmic shuttling [34]. Upon

recruitment of SR proteins for splicing, the RS domain

is hyperphosphorylated [26,28]. However, it becomes

partially dephosphorylated during splicing and this step

appears necessary for SR protein nuclear export

[46,47]. The RS domain is not sufficient, however, to

promote the export of a reporter protein. Therefore, it

regulates, rather than mediates, SR protein nuclear

export [34]. Interestingly, the non-shuttling SRSF2 is

confined to the nucleus due to the presence of a nuclear

retention sequence within its RS domain, thereby dem-

onstrating that RS domains are functionally distinct

with respect to nuclear export [45].

The shuttling ability of SR proteins is also dependent

on their RNA binding activity. This has been clearly

established for SRSF1 by earlier studies revealing that

the RNA binding activity of the first RRM is necessary

for nuclear export [34]. We recently revisited the deter-

minants of SRSF1 nuclear export by comparing the

subcellular localization of several mutants to the one of

the wild-type protein after transcription inhibition by

actinomycin D. As previously observed, this treatment

inhibits SRSF1 nuclear accumulation, leading to a

massive relocalization of SRSF1 into the cytoplasm. In

contrast, the W134A mutant in which the RNA bind-

ing capacity of RRM2 is disrupted [48] remains nuclear

under the same conditions [49]. Altogether, these data

suggest that SRSF1 nucleo-cytoplasmic shuttling

requires intact RNA binding activity of both RNA

binding domains.

Shuttling SR proteins interact with the TAP ⁄NFX1,

the primary receptor for general mRNA nuclear export

[50]. This interaction is mediated by the same domain

of TAP which is involved in the recruitment of the

Aly ⁄Ref adaptor protein. Moreover, SRSF3 and

SRSF7 were shown to activate the export of the intron-

less H2A mRNA [51]. Therefore, shuttling SR proteins

contribute to mRNA nuclear export via the interaction

with the nuclear receptor TAP. The TAP-binding

domains of SRSF1 and SRSF7 were first mapped to

the amino-terminal region composed of their RRM

and the downstream linker region [50]. Structural anal-

yses further indicated that this interaction involved

arginine residues located downstream from the RRM

of SRSF7 and SRSF3 [52] and an arginine-rich region

located between the two RNA binding domains for

SRSF1 [48]. A recent report indicates that three argi-

nine residues (R93, R97 and R109) located in SRSF1

inter-RRM linker region are methylated and strongly

influence SRSF1 nucleo-cytoplasmic distribution.

Indeed, an SRSF1 mutant in which these three arginine

residues were substituted by unmethylable and

uncharged alanine residues is predominantly located in

the cytoplasm. In contrast, a mutant in which these

arginine residues are substituted by lysines remains

predominantly nuclear, thereby suggesting that disrup-

tion of both methylation and charge of these arginine

residues modifies SRSF1 localization [32]. This study

emphasizes the importance of arginine residues located

in the TAP-binding motif for SRSF1 subcellular

distribution [48]. As hyperphosphorylated SR proteins

do not bind TAP [46,47], one could speculate that neg-

atively charged phosphoserines could associate with

these arginines, thereby blocking their interaction with

TAP. It is still to be explained, however, how the meth-

ylation status of these arginines conditions SRSF1

nucleo-cytoplasmic shuttling and how it modulates its

function as mRNA nuclear export adaptor.

In conclusion, the nuclear export of SR proteins

appears to be tightly coupled to the TAP-mediated

mRNA export pathway and to rely on a complex net-

work of sequence determinants combined with several

post-translational modifications.

Roles of SR proteins in the cytoplasm

In the last 10 years, several functions were attributed

to shuttling SR proteins in the cytoplasmic phase of

RNA metabolism. Some of these effects rely on

indirect mechanisms involving alternative splicing. For

example, high levels of SRSF1 favour the production

of an MNK2 isoform that phosphorylates the transla-

tion initiation factor eiF4E, thereby enhancing cap-

dependent translation [21]. Direct effects of SRSF1 on

cytoplasmic mRNA metabolism are diverse and

include regulation of mRNA stability and translation.

First, SR proteins modulate the NMD pathway,

whereby mRNAs containing premature termination

codons are targeted for degradation. Indeed, overex-

pression of a subset of SR proteins such as SRSF1

and SRSF2 strongly enhances NMD [53]. The NMD-

promoting effect of SRSF1 requires the presence of the

RS domain. However, this mechanism does not appear

to be dependent on SRSF1 nucleo-cytoplasmic shut-

tling, as the non-shuttling fusion protein consisting of

full-length SRSF1 fused at its C-terminus to the



nuclear retention sequence of SRSF2 retains the ability

to activate NMD. Although it remains unclear how

SR proteins activate NMD, one can hypothesize that

their recruitment promotes the nuclear modelling of

the nonsense mRNP to target it efficiently to NMD.

SRSF1 has been shown to bind to a purine-rich

sequence located in the 3¢ UTR of the mRNA encoding

a protein related to the protein-kinase-C-interacting

protein (PKCI-r) and to induce the degradation of this

transcript [54]. Furthermore, fractionation of cytosolic

extracts by sedimentation on sucrose gradients revealed

the association of SRSF1 and SRSF7 with ribosomal

subunits, monosomes and to a lesser extent polysomes,

suggesting a role for these SR proteins in mRNA

translation [55–57]. Interestingly, mutants of SRSF1

and SRSF7 lacking their RS domain are associated to

heavier polysomes than their native counterparts

[56,57]. Moreover, a hypophosphorylated SRSF1

mutant in which arginine–serine dipeptides are substi-

tuted by glycine–serines is markedly shifted to the much

heavier polysome-containing fractions, thereby suggest-

ing that RS domain hypophosphorylation favours

SRSF1 association with the translational machinery

[56]. Tethering assays indicate that SRSF1 binding to

the coding region of a reporter mRNA leads to

increased mRNA translation in cell extracts as well as

in intact cells. Moreover, while RRM1 and RS domains

are not required for this translation-enhancing effect,

RRM2 is indispensable [56]. This direct effect of SRSF1

is mediated by the recruitment of components of the

mTOR signalling pathway, resulting in phosphorylation

and release of 4E-BP, a competitive inhibitor of cap-

dependent translation [58]. This translation-activating

effect of SRSF1 is counteracted by overexpression of

SRPK2, which phosphorylates SRSF1 RS domain and

probably contributes to the dissociation of SR protein

from polysomes [56]. In contrast, it is upregulated by

mitogenic stimuli which promote SRSF1 phosphoryla-

tion by AKT kinase [30]. It thus appears that SR-

dependent translation activation is finely regulated in

response to extracellular signals and that this effect

might result from the competition between AKT- and

SRPK-mediated phosphorylation of SRSF1.

SRSF1 tethering to mRNA 3¢ UTR also activates

mRNA translation in vitro [56,58]. In intact cells, how-

ever, the opposite effect is observed [49]. The apparent

contradiction between in vitro and in vivo observations

might reflect the importance of the nuclear origin of

the target mRNA for the functional outcome of

SRSF1 recruitment to mRNA 3¢ UTR. Of note, trans-

lation inhibition mediated by SRSF1 tethering to

mRNA 3¢ UTR is markedly alleviated upon deletion

of the RS domain [49].

As mentioned above, SRSF1 controls its gene

expression partly by down-modulating the translation

of its own mRNA. This mechanism depends on the

presence of intact SRSF1 mRNA 3¢ UTR and affects

the initiation step of SRSF1 mRNA translation.

SRSF1 RRM2 is required for this regulation and

unexpectedly a nuclear-retained version of SRSF1 is

still able to autoregulate its translation. It thus appears

that nuclear SRSF1 affects the mRNP composition of

its own transcript which in turn conditions its transla-

tion efficiency in the cytoplasm [23].

SRSF1 interacts with several cellular factors includ-

ing RNA binding proteins such as TIAR and TIA-1.

These proteins are known translational repressors of

mRNAs bearing AU-rich elements (ARE) in their 3¢UTR. We have shown that overexpression of SRSF1

downregulates the expression of an ARE-containing

reporter mRNA. Interestingly, the same effect is repro-

duced upon overexpression of an RNA binding defec-

tive mutant but not upon overexpression of a mutant

lacking the RS domain [49]. Altogether, these observa-

tions suggest that SRSF1 participates in the post-tran-

scriptional control of ARE-containing mRNAs most

probably by protein–protein interactions with ARE-

binding proteins such as TIAR and TIA-1.

Finally, some viruses exploit SR proteins to promote

the translation of their mRNAs by different mecha-

nisms. SRSF3, on one hand, participates in the internal

ribosome entry site (IRES)-dependent translation of

picornavirus mRNAs. This effect is mediated by the

interaction of SRSF3 with the cellular RNA binding

protein PCBP2 that binds to IRES sequences within

viral RNAs [59]. On the other hand, SRSF5 and SRSF6

proteins activate the translation of gag protein from un-

spliced HIV-1 mRNAs. This mechanism appears to

depend on the gag-pol coding region as codon optimiza-

tion abolishes the translation-enhancing effect [60].

In conclusion, shuttling SR proteins are multifunc-

tional proteins which participate in a wide diversity of

regulatory mechanisms controlling messenger RNA

metabolism both in the nucleus and in the cytoplasm

(see Fig. 2). The regulatory output is conditioned by

multiple parameters such as the abundance of SR pro-

teins, their phosphorylation profile, their binding site

within the mRNA molecule and their association with

other factors within the mRNP.

SR proteins migrate into cytoplasmicstress granules

As they reach the cytoplasm, most mRNAs are

programmed for immediate translation, which involves

substantial RNP remodelling and assembly into



functional polysomes. However, in response to

environmental stress (heat, oxidative conditions, UV

irradiation, osmotic shock and hypoxia), eukaryotic

cells reprogramme their mRNA metabolism to repair

stress-induced damage and adapt to novel conditions.

During this process, the translation of mRNAs encod-

ing ‘housekeeping’ proteins is aborted, whereas the

translation of mRNAs encoding proteins involved in

the damage repair is stimulated. The untranslated

mRNAs concentrate in cytoplasmic foci called stress

granules (SGs) in association with stalled 48S

ribosomal pre-initiation complexes. Most of the trans-

lational machinery is recruited to SGs with the excep-

tion of the 60S subunit. In addition, SGs contain

PABP1 and several RNA-binding proteins that

regulate mRNA metabolism, including HuR, TTP,

G3BP and TIA proteins. In many cases, stress-induced

translation arrest is signalled by the phosphorylation

of translation initiator factor eIF2a. Phospho-eiF2areduces the availability of the eIF2–GTP–tRNAmet

i

ternary complex, thereby blocking translation initiation

and promoting polysome disassembly. SGs are not

stable repositories of untranslated mRNAs but rather

highly dynamic structures, as drugs that stabilize or

destabilize polysomes inhibit or promote SG assembly,

respectively, which is indicative of a dynamic equilib-

rium between these structures [61]. Moreover, upon

stress recovery, SGs are disassembled, and sequestered

mRNAs reassociate with active translational machin-

ery (see [62,63] for reviews).

On the basis of SRSF1 capacity to interact with

TIA proteins, we investigated SRSF1 migration into

SGs and found that this SR protein is recruited in

these structures in response to various stresses such as

oxidative stress, osmotic shock and heat. SRSF1 is a

bona fide SG component and does not get associated

with other cytoplasmic structures such as processing

bodies [64]. SRSF1 migration into SGs strongly

depends on its ability to bind RNA. Therefore, this

process most probably results from the sequestration

of SRSF1-bound transcripts in such cytoplasmic

structures. Accordingly, downregulation of SRSF1

expression does not alter SG assembly upon stress [49].

Other SR proteins such as SRSF2, SRSF3, SRSF7

and SRSF10 share the capacity to migrate into SGs

([65], Twyffels L. et al., unpublished results) (Table 1).

However, their propensity to assemble in such

structures is highly variable. Indeed, the shuttling

SRSF1, SRSF3 and SRSF10 are significantly relocal-

ized in cytoplasmic SGs upon stress. In contrast, the

non-shuttling SRSF2 and the shuttling SRSF7 display a

much lower propensity to assemble into SGs (Twyffels L.

et al., unpublished data). The nucleo-cytoplasmic

shuttling thus appears necessary for massive accumula-

tion of SR proteins into SGs. However, other parame-

ters seem to condition this process as shuttling SRSF7

only moderately migrates in SGs in response to oxida-

tive stress. Additionally, the fact that the non-shuttling

SRSF2 can be detected in SGs suggests that a minor

fraction of SR proteins normally retained in the

nucleus are redistributed to the cytoplasm in response

to stress. It should be noted that the nucleo-cytoplas-

mic distribution of transportin-SR is markedly altered

upon oxidative stress with significant relocalization to

the cytoplasm [66]. Therefore, the nuclear import

mediated by transportin-SR might be partially

impaired during oxidative stress, which might contrib-

ute to abnormal cytoplasmic accumulation of SR

proteins and further assembly into SGs. Altogether,

these observations suggest that the nucleo-cytoplasmic

distribution of several RNA binding proteins

undergoes major alterations upon stress and that SR

proteins participate to a variable extent in mechanisms

of translational repression ⁄derepression under these

conditions.

Conclusions

SR proteins make up a family of regulators with

important functions in RNA metabolism both in the

nucleus and in the cytoplasm. This review emphasized

the emerging roles of shuttling SR proteins in the cyto-

plasm. We have seen that SR proteins can be involved

in all phases of mRNA metabolism from synthesis to

degradation, including intermediate steps such as

nuclear export and translation. Most strikingly, the

role of SR proteins in NMD and translation regulation

perfectly exemplifies the importance of mRNP nuclear

assembly in the determination of mRNA cytoplasmic

fate and reveals some molecular links coupling gene

expression machineries.

While the parameters conditioning SRSF1 nucleo-

cytoplasmic shuttling are getting well characterized,

this is still not the case for the other shuttling SR

proteins (see Table 1). Moreover, several aspects of SR

protein distribution and activities remain unclear.

Most importantly, the signalling pathways controlling

SR domain phosphorylation as well as the phosphory-

lated residues within SR proteins are not fully identi-

fied although this post-translational modification is

known to play a major role in the control of SR

protein traffic, interactions and activities. Similarly, the

methylation of specific residues in the SRSF1 inter-

RRM region conditions SR protein localization and

thus probably modulates other aspects of SR protein

biology. Therefore, the detailed characterization of the



Table 1. Nucleo-cytoplasmic distribution, shuttling characteristics and reported functions of human SR proteins. References for the pre-

sented data are given in the text.

Name

Location

at the

equilibrium

Relocalization

to the

cytoplasm

upon RNA

PolII blockade

Shuttling in

interspecies

heterokaryon

assays

Association

with mRNA

export

machinery

Cytosolic

distribution

Accumulation

into stress

granules Reported functions

SRSF1 Nuclear

(endogenous

and tagged)

+ ++ + Free, 40S, 60S,

80S: +++

Light polysomes: +

Upon

overexpression: +

Upon various

stress: ++

Activates constitutive

and alternative splicing

Stimulates transcriptional

elongation

Contributes to genome

stability

Contributes to

mRNA export

Enhances NMD

Regulates translation

Promotes PKCI-r mRNA

degradation

Stimulates PIAS1- and

Ubc9-dependent

sumoylation

SRSF2 Nuclear

(endogenous

and tagged)

) ) ) ND Upon

overexpression: )Upon oxidative

stress: +

Upon heat

shock: )



Stimulates transcriptional

elongation

Contributes to

genome stability

Enhances NMD

SRSF3 Nuclear

(tagged)

+ ++ + Free, 40S: +++ Upon

overexpression: +

Upon oxidative

stress: ++



Contributes to mRNA export

Promotes IRES-mediated

translation of viral mRNA

SRSF4 Nuclear

(tagged)

ND + ND ND ND Activates constitutive and

alternative splicing

SRSF5 Nuclear

(tagged)

) ND ND ND ND Activates constitutive and


Stimulates translation

of unspliced HIV-1 mRNA

SRSF6 Nuclear

(endogenous

and tagged)

) + ND ND ND Activates constitutive


Stimulates translation of

unspliced HIV-1 mRNA

SRSF7 Nuclear

(tagged)

) ++ + Free: ++

40S, 60S,

80S: +++

Polysomes: +

Upon

overexpression: )Upon oxidative

stress: +

Activates constitutive and


Contributes to mRNA export

Stimulates translation

SRSF8 ND ND ND ND ND ND Regulates constitutive and


SRSF9 Nuclear

(tagged)

ND ND ND ND ND Regulates constitutive and


SRSF10 Nuclear

(tagged)

+ ++ ND Upon heat

shock: ++

Represses splicing

SRSF11 Nuclear

(endogenous)

ND ND ND ND ND Represses alternative

splicing

SRSF12 ND ND ND ND ND ND Represses alternative

splicing



post-translational modifications (e.g. phosphorylation,

methylation) of SR proteins as well as the identifica-

tion of physiological mRNA targets should provide a

more comprehensive view on how these factors

combine multiple activities in RNA metabolism. These

key issues should not wait too long to be addressed as

SR proteins exert essential cellular functions and have

been clearly established as key regulators whose unbal-

anced expression has been associated with cancer

[21,67,68] and other severe human diseases [69–71].

Acknowledgements

Laure Twyffels is an FNRS research fellow.

References

1 Zahler AM, Lane WS, Stolk JA & Roth MB (1992) SR

proteins: a conserved family of pre-mRNA splicing

factors. Genes Dev 6, 837–847.

2 Shepard PJ & Hertel KJ (2009) The SR protein family.

Genome Biol 10, 242.

3 Zhong X-Y, Wang P, Han J, Rosenfeld MG & Fu X-D

(2009) SR proteins in vertical integration of gene

expression from transcription to RNA processing to

translation. Mol Cell 35, 1–10.

4 Long JC & Caceres JF (2009) The SR protein family of

splicing factors: master regulators of gene expression.

Biochem J 417, 15–27.

5 Wu H, Sun S, Tu K, Gao Y, Xie B, Krainer AR &

Zhu J (2010) A splicing-independent function of

SF2 ⁄ASF in microRNA processing. Mol Cell 38, 66–77.

6 Boucher L, Ouzounis CA, Enright AJ & Blencowe BJ

(2001) A genome-wide survey of RS domain proteins.

RNA 7, 1693–1701.

7 Lopato S, Mayeda A, Krainer AR & Barta A (1996)

Pre-mRNA splicing in plants: characterization of

Ser ⁄Arg splicing factors. Proc Natl Acad Sci USA 93,

3074–3079.

8 Gross T, Richert K, Mierke C, Lutzelberger M &

Kaufer NF (1998) Identification and characterization of

srp1, a gene of fission yeast encoding a RNA binding

domain and a RS domain typical of SR splicing factors.

Nucleic Acids Res 26, 505–511.

9 Lutzelberger M, Gross T & Kaufer NF (1999) Srp2, an

SR protein family member of fission yeast: in vivo

characterization of its modular domains. Nucleic Acids

Res 27, 2618–2626.

10 Kress TL, Krogan NJ & Guthrie C (2008) A single

SR-like protein, Npl3, promotes pre-mRNA splicing in

budding yeast. Mol Cell 32, 727–734.

11 Manley JL & Krainer AR (2010) A rational nomencla-

ture for serine ⁄ arginine-rich protein splicing factors (SR

proteins). Genes Dev 24, 1073–1074.

12 Tacke R & Manley JL (1995) The human splicing fac-

tors ASF ⁄SF2 and SC35 possess distinct, functionally

significant RNA binding specificities. EMBO J 14,

3540–3551.

13 Fu X-D (1993) Specific commitment of different pre-

mRNAs to splicing by single SR proteins. Nature 365,

82–85.

14 Wang J & Manley JL (1995) Overexpression of the SR

proteins ASF ⁄SF2 and SC35 influences alternative splic-

ing in vivo in diverse ways. RNA 1, 335–346.

15 Das R, Yu J, Zhang Z, Gygi MP, Krainer AR, Gygi

SP & Reed R (2007) SR proteins function in coupling

RNAP II transcription to pre-mRNA splicing. Mol Cell

26, 867–881.

16 Longman D, Johnstone IL & Caceres JF (2000)

Functional characterization of SR and SR-related genes

in Caenorhabditis elegans. EMBO J 19, 1625–1637.

17 Moroy T & Heyd F (2007) The impact of alternative

splicing in vivo: mouse models show the way. RNA 13,

1155–1171.

18 Li X & Manley JL (2005) Inactivation of the SR pro-

tein splicing factor ASF ⁄SF2 results in genomic instabil-

ity. Cell 122, 365–378.

19 Pelisch F, Gerez J, Druker J, Schor IE, Munoz MJ,

Risso G, Petrillo E, Westman BJ, Lamond AI, Arzt E

et al. (2010) The serine ⁄ arginine-rich protein SF2 ⁄ASF

regulates protein sumoylation. Proc Natl Acad Sci USA

107, 116119–116124.

20 Hanamura A, Caceres JF, Mayeda A, Franza BR Jr &

Krainer AR (1998) Regulated tissue-specific expression

of antagonistic pre-mRNA splicing factors. RNA 4,

430–444.

21 Karni R, de Stanchina E, Lowe SW, Sinha R, Mu D &

Krainer AR (2007) The gene encoding the splicing fac-

tor SF2 ⁄ASF is a proto-oncogene. Nat Struct Mol Biol

14, 185–193.

22 Ni JZ, Grate L, Donohue JP, Preston C, Nobida N,

O’Brien G, Shiue L, Clark TA, Blume JE & Ares M

(2007) Ultraconserved elements are associated with

homeostatic control of splicing regulators by alternative

splicing and nonsense-mediated decay. Genes Dev 21,

708–718.

23 Sun S, Zhang Z, Sinha R, Karni R & Krainer AR

(2010) SF2 ⁄ASF autoregulation involves multiple layers

of post-transcriptional and translational control. Nat

Struct Mol Biol 17, 306–312.

24 McGlincy NJ & Smith CWJ (2008) Alternative splicing

resulting in nonsense-mediated mRNA decay: what is the

meaning of nonsense? Trends Biochem Sci 33, 385–393.

25 Meseguer S, Mudduluru G, Escamilla JM, Allgayer H

& Barettino D (2011) MicroRNAs-10a and -10b con-

tribute to retinoic acid-induced differentiation of neuro-

blastoma cells and target the alternative splicing

regulatory factor SFRS1 (SF2 ⁄ASF). J Biol Chem 286,

4150–4164.



26 Gui JF, Tronchere H, Chandler SD & Fu XD (1994)

Purification and characterization of a kinase specific for

the serine- and arginine-rich pre-mRNA splicing factors.

Proc Natl Acad Sci USA 91, 10824–10828.

27 Wang HY, Lin W, Dyck JA, Yeakley JM, Songyang Z,

Cantley LC & Fu XD (1998) SRPK2: a differentially

expressed SR protein-specific kinase involved in mediat-

ing the interaction and localization of pre-mRNA splic-

ing factors in mammalian cells. J Cell Biol 140, 737–

750.

28 Colwill K, Pawson T, Andrews B, Prasad J, Manley JL,

Bell JC & Duncan PI (1996) The Clk ⁄ Sty protein kinase

phosphorylates SR splicing factors and regulates their

intranuclear distribution. EMBO J 15, 265–275.

29 Rossi F, Labourier E, Forne T, Divita G, Derancourt

J, Riou JF, Antoine E, Cathala G, Brunel C & Tazi J

(1996) Specific phosphorylation of SR proteins by mam-

malian DNA topoisomerase I. Nature 381, 80–82.

30 Blaustein M, Pelisch F, Tanos T, Munoz MJ, Wengier

D, Quadrana L, Sanford JR, Muschietti JP, Kornblihtt

AR, Caceres JF et al. (2005) Concerted regulation of

nuclear and cytoplasmic activities of SR proteins by

AKT. Nat Struct Mol Biol 12, 1037–1044.

31 Patel NA, Kaneko S, Apostolatos HS, Bae SS, Watson

JE, Davidowitz K, Chappell DS, Birnbaum MJ, Cheng

JQ & Cooper DR (2005) Molecular and genetic studies

imply Akt-mediated signaling promotes protein kinase

CbII alternative splicing via phosphorylation of serine ⁄arginine-rich splicing factor SRp40. J Biol Chem 280,

14302–14309.

32 Sinha R, Allemand E, Zhang Z, Karni R, Myers MP &

Krainer AR (2010) Arginine methylation controls the

subcellular localization and functions of the oncoprotein

splicing factor SF2 ⁄ASF. Mol Cell Biol 30, 2762–2774.

33 Caceres JF, Misteli T, Screaton GR, Spector DL &

Krainer AR (1997) Role of the modular domains of SR

proteins in subnuclear localization and alternative splic-

ing specificity. J Cell Biol 138, 225–238.

34 Caceres JF, Screaton GR & Krainer AR (1998) A spe-

cific subset of SR proteins shuttles continuously

between the nucleus and the cytoplasm. Genes Dev 12,

55–66.

35 Zimowska G, Shi J, Munguba G, Jackson MR, Alpativ

R, Simmons MN, Shi Y & Sugrue SP (2003)

Pinin ⁄DRS ⁄memA interacts with SRp75, SRm300 and

SRrp130 in corneal epithelial cells. Invest Ophthalmol

Vis Sci 44, 4715–4723.

36 Sakashita E & Endo H (2010) SR and SR-related pro-

teins redistribute to segregated fibrillar components of

nucleoli in a response to DNA damage. Nucleus 1, 367–

380.

37 Barbe L, Lundberg E, Oksvold P, Stenius A, Lewin E,

Bjorling E, Asplund A, Ponten F, Brismar H, Uhlen M

et al. (2008) Towards a confocal subcellular atlas of the

human proteome. Mol Cell Proteomics 7, 499–508.

38 Lai M-C, Lin R-I & Tarn W-Y (2003) Differential

effects of hyperphosphorylation on splicing factor

SRp55. Biochem J 371, 937–945.

39 Cowper AE, Caceres JF, Mayeda A & Screaton GR

(2001) Serine-arginine (SR) protein-like factors that

antagonize authentic SR proteins and regulate alterna-

tive splicing. J Biol Chem 276, 48908–48914.

40 Kataoka N, Bachorik JL & Dreyfuss G (1999) Trans-

portin-SR, a nuclear import receptor for SR proteins.

J Cell Biol 145, 1145–1152.

41 Lai M-C, Lin R-I, Huang S-Y, Tsai C-W & Tarn W-Y

(2000) A human importin-b family protein, Transpor-

tin-SR2, interacts with the phosphorylated RS domain

of SR proteins. J Biol Chem 275, 7950–7957.

42 Koizumi J, Okamoto Y, Onogi H, Mayeda A, Krainer

AR & Hagiwara M (1999) The subcellular localization

of SF2 ⁄ASF is regulated by direct interaction with SR

protein kinases (SRPKs). J Biol Chem 274, 11125–11131.

43 Ngo JCK, Chakrabarti S, Ding J-H, Velazquez-Dones

A, Nolen B, Aubol BE, Adams JA, Fu X-D & Ghosh

G (2005) Interplay between SRPK and Clk ⁄Sty kinases

in phosphorylation of the splicing factor ASF ⁄SF2 is

regulated by a docking motif in ASF ⁄ SF2. Mol Cell 20,

77–89.

44 Sapra AK, Anko M-L, Grishina I, Lorenz M, Pabis M,

Poser I, Rollins J, Weiland E-M & Neugebauer KM

(2009) SR protein family members display diverse activ-

ities in the formation of nascent and mature mRNPs

in vivo. Mol Cell 34, 179–190.

45 Cazalla D, Zhu J, Manche L, Huber E, Krainer AR &

Caceres JF (2002) Nuclear export and retention signals

in the RS domain of SR proteins. Mol Cell Biol 22,

6871–6882.

46 Huang Y, Yario TA & Steitz JA (2004) A molecular

link between SR protein dephosphorylation and mRNA

export. Proc Natl Acad Sci USA 101, 9666–9670.

47 Lai M-C & Tarn W-Y (2004) Hypophosphorylated

ASF ⁄ SF2 binds TAP and is present in messenger ribo-

nucleoproteins. J Biol Chem 279, 31745–31749.

48 Tintaru AM, Hautbergue GM, Hounslow AM, Hung

M-L, Lian L-Y, Craven CJ & Wilson SA (2007) Struc-

tural and functional analysis of RNA and TAP binding

to SF2 ⁄ASF. EMBO Rep 8, 756–762.

49 Delestienne N, Wauquier C, Soin R, Dierick J-F, Guey-

dan C & Kruys V (2010) The splicing factor ASF ⁄ SF2is associated with TIA-1-related ⁄TIA-1-containing ribo-

nucleoproteic complexes and contributes to post-tran-

scriptional repression of gene expression. FEBS J 277,

2496–2514.

50 Huang Y, Gattoni R & Steitz JA (2003) SR Splicing

factors serve as adapter proteins for TAP-dependent

mRNA export. Mol Cell 11, 837–843.

51 Huang Y & Steitz JA (2001) Splicing factors SRp20

and 9G8 promote the nucleocytoplasmic export of

mRNA. Mol Cell 7, 899–905.



52 Hargous Y, Hautbergue GM, Tintaru AM, Skrisovska

L, Golovanov AP, Stevenin J, Lian L-Y, Wilson SA &

Allain FH-T (2006) Molecular basis of RNA recogni-

tion and TAP binding by the SR proteins SRp20 and

9G8. EMBO J 25, 5126–5137.

53 Zhang Z & Krainer AR (2004) Involvement of SR pro-

teins in mRNA surveillance. Mol Cell 16, 597–607.

54 Lemaire R, Prasad J, Kashima T, Gustafson J, Manley

JL & Lafyatis R (2002) Stability of a PKCI-1-related

mRNA is controlled by the splicing factor ASF ⁄ SF2:a novel function for SR proteins. Genes Dev 16, 594–607.

55 Sanford JR, Gray NK, Beckmann K & Caceres JF

(2004) A novel role for shuttling SR proteins in mRNA

translation. Genes Dev 18, 755–768.

56 Sanford JR, Ellis JD, Cazalla D & Caceres JF (2005)

Reversible phosphorylation differentially affects nuclear

and cytoplasmic functions of splicing factor 2 ⁄ alterna-tive splicing factor. Proc Natl Acad Sci USA 102,

15042–15047.

57 Swartz JE, Bor Y-C, Misawa Y, Rekosh D & Hammar-

skjold M-L (2007) The shuttling SR protein 9G8 plays

a role in translation of unspliced mRNA containing a

constitutive transport element. J Biol Chem 282, 19844–

19853.

58 Michlewski G, Sanford JR & Caceres JF (2008) The

splicing factor SF2 ⁄ASF regulates translation initiation

by enhancing phosphorylation of 4E-BP1. Mol Cell 30,

179–189.

59 Bedard KM, Daijogo S & Semler BL (2007) A nucleo-

cytoplasmic SR protein functions in viral IRES-medi-

ated translation initiation. EMBO J 26, 459–467.

60 Swanson CM, Sherer NM & Malim MH (2010) SRp40

and SRp55 promote the translation of unspliced human

immunodeficiency virus type 1 RNA. J Virol 84, 6748–

6759.

61 Kedersha N, Cho MR, Li W, Yacono PW, Chen S,

Gilks N, Golan DE & Anderson P (2000) Dynamic

shuttling of TIA-1 accompanies the recruitment of

mRNA to mammalian stress granules. J Cell Biol 151,

1257–1268.

62 Anderson P & Kedersha N (2006) RNA granules. J Cell

Biol 172, 803–808.

63 Anderson P & Kedersha N (2008) Stress granules: the

Tao of RNA triage. Trends Biochem Sci 33, 141–150.

64 Kulkarni M, Ozgur S & Stoeklin G (2010) On track

with P-bodies. Biochem Soc Trans 38, 242–251.

65 Shin C, Kleiman FE & Manley JL (2005) Multiple

properties of the splicing repressor SRp38 distinguish

it from typical SR Proteins. Mol Cell Biol 25, 8334–

8343.

66 Chang W-L & Tarn W-Y (2009) A role for Transportin

in deposition of TTP to cytoplasmic RNA granules and

mRNA decay. Nucleic Acids Res 37, 6600–6612.

67 Faustino NA & Cooper TA (2003) Pre-mRNA splicing

and human disease. Genes Dev 17, 419–437.

68 Fischer DC, Noack K, Runnebaum IB, Watermann

DO, Kieback DG, Stamm S & Stickeler E (2004)

Expression of splicing factors in human ovarian cancer.

Oncol Rep 11, 1085–1090.

69 Cartegni L & Krainer AR (2002) Disruption of an

SF2 ⁄ASF-dependent exonic splicing enhancer in SMN2

causes spinal muscular atrophy in the absence of

SMN1. Nat Genet 4, 377–384.

70 Gabut M, Mine M, Marsac C, Brivet M, Tazi J &

Soret J (2005) The SR protein SC35 is responsible for

aberrant splicing of the E1a pyruvate dehydrogenase

mRNA in a case of mental retardation with lactic

acidosis. Mol Cell Biol 25, 3286–3294.

71 Watanuki T, Funato H, Uchida S, Matsubara T,

Kobayashi A, Wakabayashi Y, Otsuki K, Nishida A &

Watanabe Y (2008) Increased expression of splicing fac-

tor SRp20 mRNA in bipolar disorder patients. J Affect

Disord 110, 62–69.



Shuttling SR proteins: more than splicing factors

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