SRSF9 promotes colorectal cancer progression via stabilizing DSN1 mRNA in an m6A-related manner

Abstract

Background Serine/arginine-rich splicing factor 9 (SRSF9) is a classical RNA-binding protein that is essential for regulating gene expression programs through its interaction with target RNA. Whether SRSF9 plays an essential role in colorectal cancer (CRC) progression and can serve as a therapeutic target is largely unknown. Here, we highlight new findings on the role of SRSF9 in CRC progression and elucidate the underlying mechanism. Methods CRC cell lines and clinical tissue samples were used. qRT-PCR, Western blotting, immunohistochemistry (IHC), gain- and loss-of-function assays, animal xenograft model studies, bioinformatic analysis, methylated single-stranded RNA affinity assays, gene-specific m6A quantitative qRT-PCR, dual-luciferase reporter assays and RNA stability assays were performed in this study. Results The expression level of SRSF9 was higher in CRC cell lines than that in an immortal human intestinal epithelial cell line. Overexpression of SRSF9 was positively associated with lymph node metastasis and Dukes stage. Functionally, SRSF9 promoted cell proliferation, migration and invasion in vitro and xenograft growth. The results of bioinformatic analysis indicated that DSN1 was the downstream target of SRSF9. In CRC cells and clinical tissue samples, the expression of SRSF9 was positively associated with the expression of DSN1. Knockdown of DSN1 partially inhibited the SRSF9-induced phenotype in CRC cells. Mechanistically, we further found that SRSF9 is an m6A-binding protein and that m6A modifications were enriched in DSN1 mRNA in CRC cells. Two m6A modification sites (chr20:36773619–36773620 and chr20:36773645–chr20:36773646) in the SRSF9-binding region (chr20:36773597–36773736) of DSN1 mRNA were identified. SRSF9 binds to DSN1 in an m6A motif- and dose-dependent manner. SRSF9 modulates the expression of DSN1 in CRC cells. Such expression regulation was largely impaired upon methyltransferase METTL3 knockdown. Moreover, knockdown of SRSF9 accelerated DSN1 mRNA turnover, while overexpression of SRSF9 stabilized DSN1 mRNA in CRC cells. Such stabilizing was also weakened upon METTL3 knockdown. Conclusion Overexpression of SRSF9 was associated with lymph node metastasis and Dukes stage in CRC. Knockdown of DSN1 eliminated the effects by SRSF9 overexpression in CRC. Our results indicated that SRSF9 functions as an m6A-binding protein (termed “reader”) by enhancing the stability of DSN1 mRNA in m6A-related manner. Our study is the first to report that SRSF9-mediated m6A recognition has a crucial role in CRC progression, and highlights SRSF9 as a potential therapeutic target for CRC management.

Differential expression profile of master regulators of gene expression- Serine-arginine rich splicing factor family in Oral Cancer

“…SRSF9 has also been implicated in cancers like hepatocellular carcinoma [22], colorectal cancer [23], and cervical cancer [24]. Frequent upregulation of SRSF10 in oral cancer has also been described earlier and it may have a role to play in its oncogenesis [25].…”

Towards understandings of serine/arginine-rich splicing factors

“…Enhanced SRSF5 phosphorylation inhibited methyltransferase-like 14 ( METTL14 ) exon10 skipping, which afterwards enhanced metastasis of pancreatic cancer cells via increasing m 6 A modification 146 . Likewise, overexpression of SRSF9 enhanced m 6 A modifications of DSN1 mRNA and was correlated with lymph node metastasis in CRC 147 .…”

ZSWIM4 inhibition improves chemosensitivity in epithelial ovarian cancer cells by suppressing intracellular glycine biosynthesis

“…Next, the slides were incubated with primary antibodies for 2 h at 37 °C, followed by treatment with a secondary antibody and diaminobenzidine. Published standards were referred to establish cutoffs for “low” and “high” ZSWIM4 expression and Ki67 scores [ 24 ]. For H&E staining, the sections were stained in hematoxylin for 5 min and eosin for 5 min.…”