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Alternative splicing of spleen tyrosine kinase differentially regulates colorectal cancer progression

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ABSTRACT

Spleen tyrosine kinase (SYK) has been reported as a potential tumor suppressor in colorectal cancer (CRC). However, the role of alternative splicing of SYK in carcinogenesis remains unclear. In the present study, SYK isoforms were overexpressed in the human CRC HCT 116 cell line using lentiviral expression vectors to investigate the biological functions of full length SYK [SYK(L)] and short form SYK [SYK(S)] in CRC. Real-time cellular analysis and the 5-ethynyl-2-deoxyuridine assay were used to detect the effects of SYK(L) and SYK(S) on cell proliferation. Cell cycle progression and migration were assessed via flow cytometry and Transwell assays, respectively. The results revealed that the recombinant lentivirus with SYK(L) overexpression significantly suppressed the proliferation and metastasis of CRC cells, while SYK(S) overexpression did not. In addition, MTS assays demonstrated that SYK(L) and SYK(S) increased the cellular sensitivity to 5-fluorouracil (5-FU), suggesting that SYK(L) and 5-FU produce a significant synergistic effect on CRC cell proliferation, while SYK(S) has an effect on modulating CRC 5-FU sensitivity. Furthermore, quantitative polymerase chain reaction results revealed that SYK(L) was downregulated in 69% of 26 pairs of CRC and adjacent non-cancerous tissues, whereas SYK(S) exhibited no significant differences between tumor and normal tissues. Overall, the present data provides evidence that SYK(L) is a tumor suppressor in CRC, and both SYK(L) and SYK(S) may serve as important predictors in the chemotherapeutic treatment of CRC.

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Human CRC HCT 116 cells transfected with recombinant lentiviral vectors with SYK(L) or SYK(S). (A) Domain structure of SYK(L) protein and its alternative splicing variant SYK(S). (B) Analysis of SYK(L) and SYK(S) expression in 7 CRC cell lines by qPCR and western blot analysis. (C) Identification of positive clones with recombinant lentiviral vectors with SYK(L) or SYK(S). (D) Detection of SYK(L) and SYK(S) expression levels in HCT 116 cells by qPCR. SYK, spleen tyrosine kinase; CRC, colorectal cancer; SYK(L), SYK full length; SYK(S), SYK short form; SH2, Src homology 2.
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f1-ol-0-0-4858: Human CRC HCT 116 cells transfected with recombinant lentiviral vectors with SYK(L) or SYK(S). (A) Domain structure of SYK(L) protein and its alternative splicing variant SYK(S). (B) Analysis of SYK(L) and SYK(S) expression in 7 CRC cell lines by qPCR and western blot analysis. (C) Identification of positive clones with recombinant lentiviral vectors with SYK(L) or SYK(S). (D) Detection of SYK(L) and SYK(S) expression levels in HCT 116 cells by qPCR. SYK, spleen tyrosine kinase; CRC, colorectal cancer; SYK(L), SYK full length; SYK(S), SYK short form; SH2, Src homology 2.

Mentions: SYK has two alternatively spliced isoforms: Full-length [SYK(L)] and short form [SYK(S)] SYK, which lacks a 69-nucleotide exon (Fig. 1A). A previous study by the present authors revealed that SYK(L) was present in the cytoplasm and nucleus of breast cancer cells, and suppressed breast cancer cell invasiveness, whereas SYK(S) was located exclusively in the cytoplasm and did not affect breast cancer cell invasion (18). Consistent with these results, recent evidence revealed that differential expression of SYK(L) and SYK(S) may contribute to tumor biology in different ways, and may be clear indicators of prognosis in patients with hepatocellular cancer (19). In addition, Prinos et al (20) have reported that changing the SYK alternative splicing pattern alters cancer cell survival and mitotic progression. On the basis of these data, the present study hypothesizes that SYK alternative splicing isoforms have different functional effects in cancer and act as modulators of cancer.


Alternative splicing of spleen tyrosine kinase differentially regulates colorectal cancer progression
Human CRC HCT 116 cells transfected with recombinant lentiviral vectors with SYK(L) or SYK(S). (A) Domain structure of SYK(L) protein and its alternative splicing variant SYK(S). (B) Analysis of SYK(L) and SYK(S) expression in 7 CRC cell lines by qPCR and western blot analysis. (C) Identification of positive clones with recombinant lentiviral vectors with SYK(L) or SYK(S). (D) Detection of SYK(L) and SYK(S) expression levels in HCT 116 cells by qPCR. SYK, spleen tyrosine kinase; CRC, colorectal cancer; SYK(L), SYK full length; SYK(S), SYK short form; SH2, Src homology 2.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4998349&req=5

f1-ol-0-0-4858: Human CRC HCT 116 cells transfected with recombinant lentiviral vectors with SYK(L) or SYK(S). (A) Domain structure of SYK(L) protein and its alternative splicing variant SYK(S). (B) Analysis of SYK(L) and SYK(S) expression in 7 CRC cell lines by qPCR and western blot analysis. (C) Identification of positive clones with recombinant lentiviral vectors with SYK(L) or SYK(S). (D) Detection of SYK(L) and SYK(S) expression levels in HCT 116 cells by qPCR. SYK, spleen tyrosine kinase; CRC, colorectal cancer; SYK(L), SYK full length; SYK(S), SYK short form; SH2, Src homology 2.
Mentions: SYK has two alternatively spliced isoforms: Full-length [SYK(L)] and short form [SYK(S)] SYK, which lacks a 69-nucleotide exon (Fig. 1A). A previous study by the present authors revealed that SYK(L) was present in the cytoplasm and nucleus of breast cancer cells, and suppressed breast cancer cell invasiveness, whereas SYK(S) was located exclusively in the cytoplasm and did not affect breast cancer cell invasion (18). Consistent with these results, recent evidence revealed that differential expression of SYK(L) and SYK(S) may contribute to tumor biology in different ways, and may be clear indicators of prognosis in patients with hepatocellular cancer (19). In addition, Prinos et al (20) have reported that changing the SYK alternative splicing pattern alters cancer cell survival and mitotic progression. On the basis of these data, the present study hypothesizes that SYK alternative splicing isoforms have different functional effects in cancer and act as modulators of cancer.

View Article: PubMed Central - PubMed

ABSTRACT

Spleen tyrosine kinase (SYK) has been reported as a potential tumor suppressor in colorectal cancer (CRC). However, the role of alternative splicing of SYK in carcinogenesis remains unclear. In the present study, SYK isoforms were overexpressed in the human CRC HCT 116 cell line using lentiviral expression vectors to investigate the biological functions of full length SYK [SYK(L)] and short form SYK [SYK(S)] in CRC. Real-time cellular analysis and the 5-ethynyl-2-deoxyuridine assay were used to detect the effects of SYK(L) and SYK(S) on cell proliferation. Cell cycle progression and migration were assessed via flow cytometry and Transwell assays, respectively. The results revealed that the recombinant lentivirus with SYK(L) overexpression significantly suppressed the proliferation and metastasis of CRC cells, while SYK(S) overexpression did not. In addition, MTS assays demonstrated that SYK(L) and SYK(S) increased the cellular sensitivity to 5-fluorouracil (5-FU), suggesting that SYK(L) and 5-FU produce a significant synergistic effect on CRC cell proliferation, while SYK(S) has an effect on modulating CRC 5-FU sensitivity. Furthermore, quantitative polymerase chain reaction results revealed that SYK(L) was downregulated in 69% of 26 pairs of CRC and adjacent non-cancerous tissues, whereas SYK(S) exhibited no significant differences between tumor and normal tissues. Overall, the present data provides evidence that SYK(L) is a tumor suppressor in CRC, and both SYK(L) and SYK(S) may serve as important predictors in the chemotherapeutic treatment of CRC.

No MeSH data available.


Related in: MedlinePlus