# RAC1B: A Rho GTPase with Versatile Functions in Malignant Transformation and Tumor Progression
## 4 Biochemical Properties, Generation and Degradation of RAC1B
### Regulation of RAC1B Splicing  
As mentioned above, RAC1B is generated from _RAC1_ by alternative RNA splicing. Using a _RAC1_ minigene in HT29 colorectal cancer (CRC) cells, [PERSON] and coworkers found that the splicing factor SRSF3 (formerly SRp20) increased skipping of alternative exon 3b, whereas another splicing factor, SRSF1 (formerly ASF/SF2), increased its inclusion [9]. RNA interference-mediated knockdown of these splicing factors confirmed that SRSF3 acts as a silencer of endogenous RAC1B splicing, whereas SRSF1 acts as an enhancer. Point mutations in exon 3b defined two adjacent regulatory regions required for skipping or inclusion of exon 3b, which are recognized in vitro by SRSF3 and SRSF1, respectively. Both splicing factors were found to be regulated by upstream signaling pathways. Activation of \(\beta\)-catenin/TCF4 increased expression of SRSF3 and hence inhibited protein levels of RAC1B while the PI3-kinase/AKT pathway increased expression of SRSF1 and promoted RAC1B generation [9].  
Alternative splicing of RAC1B via SRSF3 and SRSF1 is affected by Wnt activity in CRC cells. Wnt activity promotes expression of SRSF3, thereby blocking expression of RAC1B. Interestingly, RAC1B itself enhances Wnt signaling, forming a negative feedback loop that controls RAC1B levels through Wnt-mediated SRSF3 expression [10].  
In colorectal cells, AKT2, AKT3, GSK3B, and SR protein kinase 1 (SRPK1) increased endogenous expression of RAC1B. Knockdown of AKT2 and AKT3 affected only RAC1B protein levels suggesting a post-splicing effect, while in contrast knockdown of GSK3B or SRPK1 decreased RAC1B alternative splicing through changes in SRSF1 [11].  
[PERSON] and colleagues investigated the molecular mechanisms by which stromelysin-1/matrix metalloproteinase-3 (MMP3), a stromal enzyme upregulated in many breast tumors and other stroma-rich cancers, leads to induction of RAC1B expression. Prompted by the observation that in normal breast tissue and breast cancer biopsies, expression of the splicing factor heterogeneous nuclear ribonucleoprotein (hnRNP) A1 and RAC1B was inversely correlated, they proposed the existence of regulatory interactions between these proteins. HnRNP A1 binds to _Rac1_ exon 3b in mouse mammary epithelial cells and represses its inclusion into mature mRNA. Exposure of cells to MMP3 leads to release of hnRNP A1 from exon 3b and the consequent generation of Rac1b [12]. Importantly, hnRNP A1 (and SPSB1) have a critical role in epidermal growth factor (EGF)-driven cell migration. Mechanistically, EGF-induced ubiquitination of hnRNP A1 together with the activation of SRPKs results in the upregulation of RAC1B to promote cell motility in HeLa cells [13].  
Induction of RAC1B can also be induced by knockdown of another splicing factor, ESRP1. Downregulation of ESRP1 is closely associated with a motile phenotype of head and neck carcinoma cells. Hence, inhibition of RAC1B generation may be a potential mechanism through which ESRP1 can suppress cancer cell motility [14].