OsRhoGDI2 was isolated as a putative partner of Rho protein family member OsRacD from rice panicles by yeast two-hybrid, but its function remains unknown. In order to identify the function of OsRhoGDI2, OsRhoGDI2 knockout mutants were created by CRISPR/Cas9 technology. The results showed that two different homozygous mutants were obtained in T0 generation, and eight kinds homozygous mutants were identified in T1 generation. Sequence analysis revealed that the base substitution or base deletion occurred near the editing targets of the gene in knockout rice, and it could be expected that the truncated OsRhoGDI2 proteins lacking the RhoGDI conserved domain would be generated. Phenotype analysis showed that the OsRhoGDI2 knockout rice plants were significantly lower than the control plants. Statistical analysis confirmed that the significant decrease of plant height was due to the shortening of the second and third internodes, suggesting that OsRhoGDI2 gene may be related with rice height control.
CRISPR-Cas Systems ; Genes, Plant ; genetics ; Oryza ; genetics ; growth & development ; Plants, Genetically Modified ; rho Guanine Nucleotide Dissociation Inhibitor beta ; genetics

PURPOSE: Recent research has identified many genes and proteins that play specific roles in the process of systemic metastasis in various types of cancer. Rho GDP dissociation inhibitor 2 (RhoGDI2) has been shown to inhibit metastasis in human bladder cancer, but its role in breast cancer is controversial. MATERIALS AND METHODS: We examined the regulation and clinical significance of RhoGDI2 in Korean breast cancer patients by using proteomic approaches. RESULTS: By using a proteomic approach, we observed an increased expression of RhoGDI2 in human breast cancer tissues when compared to that of the normal breast tissues, and we validated its up-regulation in an independent cohort of 8 breast cancer patients. The clinical implication of a RhoGDI2 expression was investigated in 57 breast cancer patients by performing immunohistochemistry. RhoGDI2 did not show a significant association with the tumor size, lymph node metastasis, the histologic grade or the hormone receptor status. However, the patients with RhoGDI2-expressing tumors had significantly shorter disease-free survival (p=0.043; hazard ratio, 3.87) and distant metastasis-free survival (p=0.039; hazard ratio, 5.15). CONCLUSION: Our results demonstrated a potential role of RhoGDI2 as a poor prognostic marker as well as a potential therapeutic target. The pro-metastatic nature of RhoGDI2 shown in our study may indicate its organ-specific role in cancer metastasis.
Breast ; Breast Neoplasms ; Cohort Studies ; Disease-Free Survival ; Humans ; Immunohistochemistry ; Lymph Nodes ; Neoplasm Metastasis ; Prognosis ; Proteins ; Proteomics ; rho Guanine Nucleotide Dissociation Inhibitor beta ; Up-Regulation ; Urinary Bladder Neoplasms

OBJECTIVETo identify novel multi-drug resistance-related genes, and to explore the mechanisms of multi-drug resistance.

METHODSMulti-drug resistant cell line Lovo/5-FU was established by incubation with increasing dose of 5-FU. The sensitivity to 5-FU and cis-diaminodichloroplatinum (CDDP) was measured by MTT assay. Two dimensional electrophoresis plus mass spectrum(2-DE/MS) was used to identify the differentially expressed protein between Lovo and Lovo/5-FU. The identified protein was then verified by Western blot analysis.

RESULTSThe IC50 concentrations of Lovo/5-FU to 5-FU and CDDP were increased by 31 and 3 times, compared with Lovo (both P<0.01). 2DE-MS showed that CAP-G and RhoGDI2 were up-regulated, whereas 6-PGL, DCI, Prdx-6 and Maspin were down-regulated in Lovo/5-FU. Western blot analysis confirmed that the expression levels of RhoGDI2 and CAP-G in Lovo/5-FU were increased by 6.14 and 2.98 fold respectively (both P<0.01), whereas Maspin was decreased to 5.2% of Lovo(P<0.01).

CONCLUSIONSMulti-gene and multi-pathway are involved in the development of multi-drug resistance of colorectal cancer cells. CAP-G, RhoGDI2 and Maspin are potential multi-drug resistant genes.

Cell Line, Tumor ; Colonic Neoplasms ; genetics ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Humans ; Microfilament Proteins ; genetics ; Nuclear Proteins ; genetics ; Serpins ; genetics ; rho Guanine Nucleotide Dissociation Inhibitor beta ; genetics