Sik (BRK) Phosphorylates Sam68 in the Nucleus and Negatively Regulates Its RNA Binding Ability

Abstract

Sik (mouse Src-related intestinal kinase) and its orthologue BRK (human breast tumor kinase) are intracellular tyrosine kinases that are distantly related to the Src family and have a similar structure, but they lack the myristoylation signal. Here we demonstrate that Sik and BRK associate with the RNA binding protein Sam68 (Src associated during mitosis, 68 kDa). We found that Sik interacts with Sam68 through its SH3 and SH2 domains and that the proline-rich P3 region of Sam68 is required for Sik and BRK SH3 binding. In the transformed HT29 adenocarcinoma cell cell line, endogenous BRK and Sam68 colocalize in Sam68-SLM nuclear bodies (SNBs), while transfected Sik and Sam68 are localized diffusely in the nucleoplasm of nontransformed NMuMG mammary epithelial cells. Transfected Sik phosphorylates Sam68 in SNBs in HT29 cells and in the nucleoplasm of NMuMG cells. In functional studies, expression of Sik abolished the ability of Sam68 to bind RNA and act as a cellular Rev homologue. While Sam68 is a substrate for Src family kinases during mitosis, Sik/BRK is the first identified tyrosine kinase that can phosphorylate Sam68 and regulate its activity within the nucleus, where it resides during most of the cell cycle.

Breast Tumor Kinase (BRK) Signaling in Breast Cancer

“…In breast cancer cell lines, BRK is present in SNBs where it associates with and can phosphorylate Sam68 (Deny et al, 2000) Although the biological function of Sam68 is unknown, Sam68 can function as a cellular homologue of Rev by transporting unspliced HIV mRNA into the cytoplasm (Reddy et al, 1999). In our studies, we found that phosphorylation of Sam68 by BRK/Sik negatively regulates its RNA binding ability and its ability to function as a Rev cellular homologue (Derry et al, 2000). These data strengthen the possibility that the Recently, two Sam68-like-mammalian proteins, SLM-1 and SLM-2 were identified (Di Fruscio et al, 1999).…”

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

“…In recent years, several additional PTK6 substrates have been identified (Derry et al, 2000;Babic et al, 2004;Haegebarth et al, 2004Haegebarth et al, , 2005. The first reported substrate of PTK6 phosphorylation was Sam68 (Src-associated in mitosis 68 kDa), and it was shown that PTK6 negatively regulates its RNA-binding activity (Derry et al, 2000).…”

“…The first reported substrate of PTK6 phosphorylation was Sam68 (Src-associated in mitosis 68 kDa), and it was shown that PTK6 negatively regulates its RNA-binding activity (Derry et al, 2000). This may have an impact on the post-transcriptional regulation of gene expression.…”

Interactions of STAP-2 with BRK and STAT3/5 in Breast Cancer Cells

“…Sam68 is the most extensively studied BRK substrate. BRK expression suppressed cell proliferation through EGFR-mediated phosphorylation of Sam68 in a human breast cancer cell line (Coyle et al, 2003;Derry et al, 2000;Lukong et al, 2005). Similarly, BRK phosphorylates the Sam68-like mammalian proteins, SLM-1 and SLM-2, and negatively regulates their RNA-binding functions (Haegebarth et al, 2004).…”

“…BRK substrates include RNA-binding proteins (Sam68 (Coyle et al, 2003;Derry et al, 2000;Lukong et al, 2005), SLM-1/2 (Haegebarth et al, 2004), and the polypyrimidine tractbinding protein-associated splicing factor (PSF) (Lukong et al, 2009)), transcription factors (STAT3 (Liu et al, 2006) and STAT5A/B (Weaver and Silva, 2007)), adaptor molecules (STAP-2) (Mitchell et al, 2000), and a variety of signaling molecules (paxillin (Chen et al, 2004), p190RhoGAP (Shen et al, 2008), kinesin-associated protein 3A (KAP3A) (Lukong and Richard, 2008), Akt (Zhang et al, 2005), -catenin (Palka-Hamblin et al, 2010), and ARAP1 (Arf-GAP, Rho-GAP, ankyrin repeat and PH domain-containing protein 1; also known as centaurin δ-2) (Kang et al, 2010)). Although BRK expression is known to induce tyrosine phosphorylation in some of these, similar actions in others have yet to be confirmed.…”