Sam68 Proteins Research Articles

The SWI/SNF subunit BRG1 affects alternative splicing by changing RNA binding factor interactions with nascent RNA

BRG1 and BRM are ATPase core subunits of the human SWI/SNF chromatin remodelling complexes mainly associated with transcriptional initiation. They also have a role in alternative splicing, which has been shown for BRM-containing SWI/SNF complexes at a few genes. Here, we have identified a subset of genes which harbour alternative exons that are affected by SWI/SNF ATPases by expressing the ATPases BRG1 and BRM in C33A cells, a BRG1- and BRM-deficient cell line, and analysed the effect on splicing by RNA sequencing. BRG1- and BRM-affected sub-sets of genes favouring both exon inclusion and exon skipping, with only a minor overlap between the ATPase. Some of the changes in alternative splicing induced by BRG1 and BRM expression did not require the ATPase activity. The BRG1-ATPase independent included exons displayed an exon signature of a high GC content. By investigating three genes with exons affected by the BRG-ATPase-deficient variant, we show that these exons accumulated phosphorylated RNA pol II CTD, both serine 2 and serine 5 phosphorylation, without an enrichment of the RNA polymerase II. The ATPases were recruited to the alternative exons, together with both core and signature subunits of SWI/SNF complexes, and promoted the binding of RNA binding factors to chromatin and RNA at the alternative exons. The interaction with the nascent RNP, however, did not reflect the association to chromatin. The hnRNPL, hnRNPU and SAM68 proteins associated with chromatin in cells expressing BRG1 and BRM wild type, but the binding of hnRNPU to the nascent RNP was excluded. This suggests that SWI/SNF can regulate alternative splicing by interacting with splicing-RNA binding factor and influence their binding to the nascent pre-mRNA particle.

The Oncogene Metadherin Interacts with the Known Splicing Proteins YTHDC1, Sam68 and T-STAR and Plays a Novel Role in Alternative mRNA Splicing.

Oncogenic metadherin is a key contributor to tumourigenesis with metadherin expression and cytoplasmic localisation previously linked to poor survival. A number of reports have shown metadherin localises specifically to nuclear speckles known to be rich in RNA-binding proteins including the splicing proteins YTHDC1, Sam68 and T-STAR, that have been shown to select alternative splice sites in mRNA of tumour-associated proteins including BRCA, MDM2 and VEGF. Here we investigate the interaction and relationship between metadherin and the splice factors YTHDC1, T-STAR and Sam68. Using a yeast two-hybrid assay and immunoprecipitation we show that metadherin interacts with YTHDC1, Sam68 and T-STAR and demonstrate that T-STAR is significantly overexpressed in prostate cancer tissue compared to benign prostate tissue. We also demonstrate that metadherin influences splice site selection in a dose-dependent manner in CD44v5-luc minigene reporter assays. Finally, we demonstrate that prostate cancer patients with higher metadherin expression have greater expression of the CD44v5 exon. CD44v5 expression could be used to discriminate patients with poor outcomes following radical prostatectomy. In this work we show for the first time that metadherin interacts with, and modulates, the function of key components of splicing associated with cancer development and progression.

Binding site density enables paralog-specific activity of SLM2 and Sam68 proteins in Neurexin2 AS4 splicing control.

SLM2 and Sam68 are splicing regulator paralogs that usually overlap in function, yet only SLM2 and not Sam68 controls the Neurexin2 AS4 exon important for brain function. Herein we find that SLM2 and Sam68 similarly bind to Neurexin2 pre-mRNA, both within the mouse cortex and in vitro. Protein domain-swap experiments identify a region including the STAR domain that differentiates SLM2 and Sam68 activity in splicing target selection, and confirm that this is not established via the variant amino acids involved in RNA contact. However, far fewer SLM2 and Sam68 RNA binding sites flank the Neurexin2 AS4 exon, compared with those flanking the Neurexin1 and Neurexin3 AS4 exons under joint control by both Sam68 and SLM2. Doubling binding site numbers switched paralog sensitivity, by placing the Neurexin2 AS4 exon under joint splicing control by both Sam68 and SLM2. Our data support a model where the density of shared RNA binding sites around a target exon, rather than different paralog-specific protein–RNA binding sites, controls functional target specificity between SLM2 and Sam68 on the Neurexin2 AS4 exon. Similar models might explain differential control by other splicing regulators within families of paralogs with indistinguishable RNA binding sites.

Structural basis of RNA recognition and dimerization by the STAR proteins T-STAR and Sam68.

Sam68 and T-STAR are members of the STAR family of proteins that directly link signal transduction with post-transcriptional gene regulation. Sam68 controls the alternative splicing of many oncogenic proteins. T-STAR is a tissue-specific paralogue that regulates the alternative splicing of neuronal pre-mRNAs. STAR proteins differ from most splicing factors, in that they contain a single RNA-binding domain. Their specificity of RNA recognition is thought to arise from their property to homodimerize, but how dimerization influences their function remains unknown. Here, we establish at atomic resolution how T-STAR and Sam68 bind to RNA, revealing an unexpected mode of dimerization different from other members of the STAR family. We further demonstrate that this unique dimerization interface is crucial for their biological activity in splicing regulation, and suggest that the increased RNA affinity through dimer formation is a crucial parameter enabling these proteins to select their functional targets within the transcriptome.

Up-Regulation of the RNA-Binding Proteins SAM68 and PTB Contributes to the Acquisition of a Drug-Resistance Phenotype in Pancreatic Adenocarcinoma Cells

Context The limited effect of conventional chemotherapy in pancreatic adenocarcinoma (PDAC) urges for novel therapies, targeting more directly the molecular aberrations of this disease. The molecular characterization of the drug resistant phenotype of PDAC cells remain largely unexplored, even though some evidence suggests a correlation with the expression of mesenchymal markers. Notably, the epithelial-to-mesenchymal transition (EMT) is promoted by finely-tuned changes in gene expression, occurring at both transcriptional and splicing levels. In this regard, recent observations have shown the requirement for select splicing factors during EMT. Objective Characterization of the molecular events leading to drug resistance in PDAC cells. Methods Chronic exposure to gemcitabine to select drug-resistant PDAC cell subpopulations. Western blot analyses for the expression of cancer related proteins, RNA-interference of selected genes to investigate their function. Trypan blue cell count, immuno­fluorescence analysis of the cleaved of caspase-3 and clonogenic assay to analyze cell survival. PCR analysis to detect EMT markers and immuno­fluorescence analysis of phalloidin to detect actin cytoskeleton morphology. Scratch assay to test cell migration. Results The chronic exposure of PDAC cells to gemcitabine selected a subpopulation of cells that displays a mesenchymal phenotype and is less sensitive to drug-induced cell death. These cells express higher levels of SAM68 and PTB, two splicing factors with oncogenic functions in other types of human cancers. Depletion of SAM68 and PTB expression caused a partial recovery of drug sensitivity, suggesting that they contribute to the acquisition of the drug-resistant phenotype. Conclusion Our data indicate that SAM68 and PTB may represent suitable molecular targets to counteract the drug resistance of PDAC cells.

Involvement of the RNA-binding proteins Sam68 and PTB in the acquisition of the resistance to gemcitabine in pancreatic adenocarcinoma cells

Involvement of the RNA-binding proteins Sam68 and PTB in the acquisition of the resistance to gemcitabine in pancreatic adenocarcinoma cells

The Tissue-Specific RNA Binding Protein T-STAR Controls Regional Splicing Patterns of Neurexin Pre-mRNAs in the Brain

The RNA binding protein T-STAR was created following a gene triplication 520–610 million years ago, which also produced its two parologs Sam68 and SLM-1. Here we have created a T-STAR null mouse to identify the endogenous functions of this RNA binding protein. Mice null for T-STAR developed normally and were fertile, surprisingly, given the high expression of T-STAR in the testis and the brain, and the known infertility and pleiotropic defects of Sam68 null mice. Using a transcriptome-wide search for splicing targets in the adult brain, we identified T-STAR protein as a potent splicing repressor of the alternatively spliced segment 4 (AS4) exons from each of the Neurexin1-3 genes, and exon 23 of the Stxbp5l gene. T-STAR protein was most highly concentrated in forebrain-derived structures like the hippocampus, which also showed maximal Neurexin1-3 AS4 splicing repression. In the absence of endogenous T-STAR protein, Nrxn1-3 AS4 splicing repression dramatically decreased, despite physiological co-expression of Sam68. In transfected cells Neurexin3 AS4 alternative splicing was regulated by either T-STAR or Sam68 proteins. In contrast, Neurexin2 AS4 splicing was only regulated by T-STAR, through a UWAA-rich response element immediately downstream of the regulated exon conserved since the radiation of bony vertebrates. The AS4 exons in the Nrxn1 and Nrxn3 genes were also associated with distinct patterns of conserved UWAA repeats. Consistent with an ancient mechanism of splicing control, human T-STAR protein was able to repress splicing inclusion of the zebrafish Nrxn3 AS4 exon. Although Neurexin1-3 and Stxbp5l encode critical synaptic proteins, T-STAR null mice had no detectable spatial memory deficits, despite an almost complete absence of AS4 splicing repression in the hippocampus. Our work identifies T-STAR as an ancient and potent tissue-specific splicing regulator that uses a concentration-dependent mechanism to co-ordinately regulate regional splicing patterns of the Neurexin1-3 AS4 exons in the mouse brain.

Involvement of the RNA-binding proteins Sam68 and SRSF1 in the acquisition of the resistance to gemcitabine in pancreatic adenocarcinoma cells

Involvement of the RNA-binding proteins Sam68 and SRSF1 in the acquisition of the resistance to gemcitabine in pancreatic adenocarcinoma cells

Involvement of the RNA-binding proteins Sam68 and SRSF1 in the acquisition of the resistance to gemcitabine in pancreatic adenocarcinoma cells

Involvement of the RNA-binding proteins Sam68 and SRSF1 in the acquisition of the resistance to gemcitabine in pancreatic adenocarcinoma cells

The STAR family proteins Sam68, Slm-1 and Slm-2 differentially regulate proliferation and differentiation of cortical neural stem/progenitor cells

The STAR family proteins Sam68, Slm-1 and Slm-2 differentially regulate proliferation and differentiation of cortical neural stem/progenitor cells

Structural Basis for Homodimerization of the Src-associated during Mitosis, 68-kDa Protein (Sam68) Qua1 Domain

Sam68 (Src-associated during mitosis, 68 kDa) is a prototypical member of the STAR (signal transducer and activator of RNA) family of RNA-binding proteins. STAR proteins bind mRNA targets and modulate cellular processes such as cell cycle regulation and tissue development in response to extracellular signals. Sam68 has been shown to modulate alternative splicing of the pre-mRNAs of CD44 and Bcl-xL, which are linked to tumor progression and apoptosis. Sam68 and other STAR proteins recognize bipartite RNA sequences and are thought to function as homodimers. However, the structural and functional roles of the self-association are not known. Here, we present the solution structure of the Sam68 Qua1 homodimerization domain. Each monomer consists of two antiparallel alpha-helices connected by a short loop. The two subunits are arranged perpendicular to each other in an unusual four-helix topology. Mutational analysis of Sam68 in vitro and in a cell-based assay revealed that the Qua1 domain and residues within the dimerization interface are essential for alternative splicing of a CD44 minigene. Together, our results indicate that the Qua1 homodimerization domain is required for regulation of alternative splicing by Sam68.

Proteomic identification of heterogeneous nuclear ribonucleoprotein L as a novel component of SLM/Sam68 Nuclear Bodies

BackgroundActive pre-mRNA splicing occurs co-transcriptionally, and takes place throughout the nucleoplasm of eukaryotic cells. Splicing decisions are controlled by networks of nuclear RNA-binding proteins and their target sequences, sometimes in response to signalling pathways. Sam68 (Src-associated in mitosis 68 kDa) is the prototypic member of the STAR (Signal Transduction and Activation of RNA) family of RNA-binding proteins, which regulate splicing in response to signalling cascades. Nuclear Sam68 protein is concentrated within subnuclear organelles called SLM/Sam68 Nuclear Bodies (SNBs), which also contain some other splicing regulators, signalling components and nucleic acids.ResultsWe used proteomics to search for the major interacting protein partners of nuclear Sam68. In addition to Sam68 itself and known Sam68-associated proteins (heterogeneous nuclear ribonucleoproteins hnRNP A1, A2/B1 and G), we identified hnRNP L as a novel Sam68-interacting protein partner. hnRNP L protein was predominantly present within small nuclear protein complexes approximating to the expected size of monomers and dimers, and was quantitatively associated with nucleic acids. hnRNP L spatially co-localised with Sam68 as a novel component of SNBs and was also observed within the general nucleoplasm. Localisation within SNBs was highly specific to hnRNP L and was not shared by the closely-related hnRNP LL protein, nor any of the other Sam68-interacting proteins we identified by proteomics. The interaction between Sam68 and hnRNP L proteins was observed in a cell line which exhibits low frequency of SNBs suggesting that this association also takes place outside SNBs. Although ectopic expression of hnRNP L and Sam68 proteins independently affected splicing of CD44 variable exon v5 and TJP1 exon 20 minigenes, these proteins did not, however, co-operate with each other in splicing regulation of these target exons.ConclusionHere we identify hnRNP L as a novel SNB component. We show that, compared with other identified Sam68-associated hnRNP proteins and hnRNP LL, this co-localisation within SNBs is specific to hnRNP L. Our data suggest that the novel Sam68-hnRNP L protein interaction may have a distinct role within SNBs.

The STAR RNA binding proteins GLD-1, QKI, SAM68 and SLM-2 bind bipartite RNA motifs

BackgroundSAM68, SAM68-like mammalian protein 1 (SLM-1) and 2 (SLM-2) are members of the K homology (KH) and STAR (signal transduction activator of RNA metabolism) protein family. The function of these RNA binding proteins has been difficult to elucidate mainly because of lack of genetic data providing insights about their physiological RNA targets. In comparison, genetic studies in mice and C. elegans have provided evidence as to the physiological mRNA targets of QUAKING and GLD-1 proteins, two other members of the STAR protein family. The GLD-1 binding site is defined as a hexanucleotide sequence (NACUCA) that is found in many, but not all, physiological GLD-1 mRNA targets. Previously by using Systematic Evolution of Ligands by EXponential enrichment (SELEX), we defined the QUAKING binding site as a hexanucleotide sequence with an additional half-site (UAAY). This sequence was identified in QKI mRNA targets including the mRNAs for myelin basic proteins.ResultsHerein we report using SELEX the identification of the SLM-2 RNA binding site as direct U(U/A)AA repeats. The bipartite nature of the consensus sequence was essential for SLM-2 high affinity RNA binding. The identification of a bipartite mRNA binding site for QKI and now SLM-2 prompted us to determine whether SAM68 and GLD-1 also bind bipartite direct repeats. Indeed SAM68 bound the SLM-2 consensus and required both U(U/A)AA motifs. We also confirmed that GLD-1 also binds a bipartite RNA sequence in vitro with a short RNA sequence from its tra-2 physiological mRNA target.ConclusionThese data demonstrate that the STAR proteins QKI, GLD-1, SAM68 and SLM-2 recognize RNA with direct repeats as bipartite motifs. This information should help identify binding sites within physiological RNA targets.

Prognostic value of protein tyrosine kinase 6 (PTK6) for long-term survival of breast cancer patients

The cytoplasmic tyrosine kinase PTK6 (BRK) shows elevated expression in approximately two-thirds of primary breast tumours, and is implicated in EGF receptor-dependent signalling and epithelial tumorigenesis. Using immunohistochemistry, we performed a retrospective study on 426 archival breast cancer samples from patients with long-term follow-up and compared the protein expression levels of PTK6, the HER receptors, Sam68 (a substrate of PTK6), and signalling proteins including MAP kinase (MAPK), phosphorylated MAPK (P-MAPK), and PTEN. We show that PTK6 expression is of significant prognostic value in the outcome of breast carcinomas. In multivariate analysis, the disease-free survival of patients of ⩾240 months was directly associated with the protein expression level of PTK6 (P⩽0.001), but was also inversely associated with nodal status (P⩽0.001) and tumour size (P⩽0.01). PTK6 expression in tumour tissue significantly correlated (P⩽0.05) with the expression of PTEN, MAPK, P-MAPK, and Sam68. To investigate whether these correlations may be due to molecular interactions between PTK6 and these proteins, we used protein extracts from the T47D cell line for immunoprecipitation and western blot analysis. By this, interactions could be demonstrated between PTK6 and MAPK, P-MAPK, HER2/neu, HER3, HER4, PTEN, and Sam68. On the basis of these results, we suggest that PTK6 may serve as a future target for the development of novel treatments in breast cancer.

Promiscuous Modification of the Nuclear Poly(A)-binding Protein by Multiple Protein-arginine Methyltransferases Does Not Affect the Aggregation Behavior

The mammalian nuclear poly(A)-binding protein, PABPN1, carries 13 asymmetrically dimethylated arginine residues in its C-terminal domain. By fractionation of cell extracts, we found that protein-arginine methyltransferases (PRMTs)-1, -3, and -6 are responsible for the modification of PABPN1. Recombinant PRMT1, -3, and -6 also methylated PABPN1. Our data suggest that these enzymes act on their own, and additional polypeptides are not involved in recognizing PABPN1 as a substrate. PRMT1 is the predominant methyltransferase acting on PABPN1. Nevertheless, PABPN1 was almost fully methylated in a Prmt1(-/-) cell line; thus, PRMT3 and -6 suffice for methylation. In contrast to PABPN1, the heterogeneous nuclear ribonucleoprotein (hnRNP) K is selectively methylated only by PRMT1. Efficient methylation of synthetic peptides derived from PABPN1 or hnRNP K suggested that PRMT1, -3, and -6 recognize their substrates by interacting with local amino acid sequences and not with additional domains of the substrates. However, the use of fusion proteins suggested that the inability of PRMT3 and -6 to modify hnRNP K is because of structural masking of the methyl-accepting amino acid sequences by neighboring domains. Mutations leading to intracellular aggregation of PABPN1 cause the disease oculopharyngeal muscular dystrophy. The C-terminal domain containing the methylated arginine residues is known to promote PAPBN1 self-association, and arginine methylation has been reported to inhibit self-association of an orthologous protein. Thus, arginine methylation might be relevant for oculopharyngeal muscular dystrophy. However, in two different types of assays we have been unable to detect any effect of arginine methylation on the aggregation of bovine PABPN1.

Bcl-xS Is an Important Pro-Apoptotic Bcl-2 Family Member in Chronic Lymphocytic Leukaemia and Its Expression Is Regulated by SAM68, hnRNP-F and hnRNP-H.

We have previously shown that the pro-apoptotic Bcl-x isoform Bcl-xS is induced in chronic lymphocytic leukaemia cells during spontaneous and drug induced apoptosis to a greater extent than other pro-apoptotic Bcl-2 family members including Bim, Puma and Noxa. Puma, Noxa and also Bid were induced strongly by stimulation with CD40 ligand, in culture conditions causing CLL cell proliferation and survival, and associated with strong induction of the pro-survival Bcl-x isoform, Bcl-xL. Immunohistochemistry demonstrates Bcl-xL expression in lymph node proliferation centres. We, therefore, conclude that splicing of Bcl-x mRNA is important for regulation of CLL survival such that Bcl-xL is required during proliferation in the lymph node or bone marrow microenvironment, but Bcl-xS is induced during apoptosis. Splicing of Bcl-x mRNA in HeLa cells is known to be controlled by RNA binding proteins including SAM68, heterogeneous ribonucleoprotein-F (hnRNP-F) and hnRNP-H, but expression and regulation of these molecules has not been investigated in CLL. We have demonstrated that expression of hnRNP-F and hnRNP-H decline in association with an increase in Bcl-xS expression. Conversely expression of the ribonucleoproteins is maintained (relative to expression in freshly isolated peripheral blood CLL cells) or increases on CD40 stimulation. SAM68 showed a similar pattern of expression with increased expression on CD40 stimulation and decreased under conditions of apoptosis. Phosphorylation of SAM68 can control the RNA binding activity of this molecule. We immunoprecipitated SAM68 from freshly isolated CLL cells and cells that had been cultured with CD40 ligand (to promote proliferation) or on tissue culture plastic (to promote apoptosis) for 2 days. Westerns of the immunoprecipitated protein were probed with an anti-phosphotyrosine antibody. We found variable patterns: phosphorylation was uniformly present in cells cultured on plastic (4/4 cases) but was only detectable in freshly isolated cells in 1 case and on CD40 stimulation in 2 cases. We conclude that the RNA binding proteins hnRNP-F, hnRNP-H and SAM68 have important roles in controlling Bcl-x splicing in CLL cells, and that their effects are largely due to variation in expression level rather than phosphorylation. Strategies to bias production of pro-apoptotic Bcl-xS using siRNA either directed against Bcl-xL mRNA or against RNA binding proteins may be therapeutically useful.

Arginine methylation of Sam68 and SLM proteins negatively regulates their poly(U) RNA binding activity

Sam68 ( Src substrate associated during mitosis) and its homologues, SLM-1 and SLM-2 ( Sam68- like mammalian proteins), are RNA binding proteins and contain the arg-gly (RG) repeats, in which arginine residues are methylated by the protein arginine methyltransferase 1 (PRMT1). However, it remains unclear whether the arginine methylation affects an RNA binding. Here, we report that methylation of Sam68 and SLM proteins markedly reduced their poly(U) binding ability in vitro. The RG repeats of Sam68 bound poly(U), but arginine methylation of the RG repeats abrogated its poly(U) binding ability in vitro. Overexpression of PRMT1 increased arginine methylation of Sam68 and SLM proteins in cells, which resulted in a decrease of their poly(U) binding ability. The results suggest that the RG repeats conserved in Sam68 and SLM proteins may function as an auxiliary RNA binding domain and arginine methylation may eliminate or reduce an RNA binding ability of the proteins.

Alternative RNA splicing complexes containing the scaffold attachment factor SAFB2

The scaffold attachment factor SAFB1 and its recently discovered homologue SAFB2 might provide an important link between pre-mRNA splicing, intracellular signalling and transcription. Using novel mono-specific antisera, we found endogenous SAFB2 protein has a different spatial distribution from SAFB1 within the nucleus where it is found in much larger nuclear complexes (up to 670 kDa in size), and a distinct pattern of expression in adult human testis. By contrast, SAFB1 protein predominantly exists either as smaller complexes or as a monomeric protein. Our results suggest stable core complexes containing components comprised of SAFB1, SAFB2 and the RNA binding proteins Sam68 and hnRNPG exist in parallel with free SAFB1 protein. We found that SAFB2 protein, like SAFB1, acts as a negative regulator of a tra2beta variable exon. Despite showing an involvement in splicing, we detected no stable interaction between SAFB proteins and SR or SR-related splicing regulators, although these were also found in stable higher molecular mass complexes. Each of the detected alternative splicing regulator complexes exists independently of intact nucleic acids, suggesting they might be pre-assembled and recruited to nascent transcripts as modules to facilitate alternative splicing, and/or they represent nuclear storage compartments from which active proteins are recruited.

Measles virus induces expression of SIP110, a constitutively membrane clustered lipid phosphatase, which inhibits T cell proliferation

Interference of measles virus (MV) with phosphatidyl-inositol-3-kinase (PI3K) activation in response to T cell receptor ligation was identified as important for the induction of T cell paralysis. We now show that MV exposure of unstimulated T cells induces expression of SIP110, an isoform of the lipid phosphatase SHIP145, which is translated from an intron-derived sequences containing mRNA. We found that MV contact can regulate stimulated exon inclusion into pre-mRNAs by targeting PI3K or MAPK-dependent nuclear translocation and activation of splicing regulatory serine-arginine rich (SR) and Sam68 proteins. Induction of SIP110 in resting T cells relied on MV-dependent interference with basal activity of the PI3K. SIP110 was cloned from MV-exposed T cells, and, when transiently expressed in primary or Jurkat T cells, localized into membrane clusters independently of T cell activation. Confirming that SIP110 is a catalytically active lipid phosphatase, its transgenic expression abolished basal and impaired PMA/ionomycin-stimulated phosphorylation of the Akt kinase which is important for T cell proliferation. Thus MV causes induction of SIP110 expression, which constitutively depletes the cellular phosphoinositol-3,4,5-phosphate pool suggesting that thereby the threshold for activation signals necessary for the induction of T cell proliferation is raised.

The Nuclear Tyrosine Kinase BRK/Sik Phosphorylates and Inhibits the RNA-binding Activities of the Sam68-like Mammalian Proteins SLM-1 and SLM-2

Expression of the intracellular tyrosine kinase BRK/Sik is epithelial-specific and regulated during differentiation. Only a few substrates have been identified for BRK/Sik, including the KH domain containing RNA-binding protein Sam68 and the novel adaptor protein BKS. Although the physiological role of Sam68 is unknown, it has been shown to regulate mRNA transport, pre-mRNA splicing, and polyadenylation. Here we demonstrate that the Sam68-like mammalian proteins SLM-1 and SLM-2 but not the related KH domain containing heterogeneous nuclear ribonucleoprotein K are novel substrates of BRK/Sik. The expression of active BRK/Sik results in increased SLM-1 and SLM-2 phosphorylation and increased retention of BRK/Sik within the nucleus. The phosphorylation of SLM-1 and SLM-2 has functional relevance and leads to inhibition of their RNA-binding abilities. We show that SLM-1, SLM-2, and BRK/Sik have restricted patterns of expression unlike the ubiquitously expressed Sam68. Moreover, BRK/Sik, SLM-1, and Sam68 transcripts were coexpressed in the mouse gastrointestinal tract and skin, suggesting that SLM-1 and Sam68 could be physiologically relevant BRK/Sik targets in vivo. The ability of BRK/Sik to negatively regulate the RNA-binding activities of the KH domain RNA binding proteins SLM-1 and Sam68 may have an impact on the posttranscriptional regulation of epithelial cell gene expression.