Nectin-like molecule 2, a necessary sexual maturation regulator, participates in congenital hypogonadotropic hypogonadism.

BACKGROUND: Congenital hypogonadotropic hypogonadism (CHH) is a rare genetically heterogeneous disorder. We aimed to determine the prevalence and pathogenesis of NECL2 (Nectin-like molecule 2) variants in a cohort of female patients with CHH. METHODS: We sequenced and determined the prevalence of NECL2 variants in 68 female patients with CHH and 243 healthy controls collected from an academic medical center. Further cellular and animal studies were performed to verify the pathogenicity of the mutations. Necl2 knockout female mice were generated, and their puberty development was observed. RESULTS: A novel NECL2 variant (c.1052_1060del, p.Thr351_Thr353del) was detected in 4 of 68 (5.9%) patients with CHH. Its prevalence was significantly higher in CHH patients than in healthy controls (0%). At the cellular level, the necl2 variant leads to a decrease in gonadotropin-releasing hormone. In animal models, we found that the Necl2 protein was expressed in the hypothalamus, especially in the ventromedial hypothalamic nucleus of mice. Necl2 knockout female mice showed delayed puberty and an irregular estrous cycle, consistent with CHH patient phenotypes. CONCLUSIONS: Our findings predict that NECL2 may be a new candidate gene for CHH and that the NECL2 protein plays a critical role in the progression of puberty development.

Structure of the heterophilic interaction between the nectin-like 4 and nectin-like 1 molecules.

Nectin-like (Necl) molecules are Ca2+-independent Ig-like transmembrane cell adhesion molecules that participate in junctions between different cell types. The specific cell-cell adhesions mediated by Necl proteins are important in neural development and have been implicated in neurodegenerative diseases. Here, we present the crystal structure of the mouse Necl-4 full ectodomain and the structure of the heterophilic Necl ectodomain complex formed by the mNecl-4 and mNecl-1 ectodomains. We demonstrate that, while the ectodomain of mNecl-4 is monomeric, it forms a stable heterodimer with Ig1 of mNecl-1, with an affinity significantly higher than that observed for self-dimerization of the mNecl-1 ectodomain. We validated our structural characterizations by performing a surface plasmon resonance assay and an Fc fusion protein binding assay in mouse primary dorsal root ganglia neurites and Schwann cells and identified a selection of residues important for heterophilic interactions. Finally, we proposed a model of Necl binding specificity that involves an induced-fit conformational change at the dimerization interface.

High expression of PTBP1 promote invasion of colorectal cancer by alternative splicing of cortactin.

Polypyrimidine tract-binding protein 1 (PTBP1) involving in almost all steps of mRNA regulation including alternative splicing metabolism during tumorigenesis due to its RNA-binding activity. Initially, we found that high expressed PTBP1 and poor prognosis was interrelated in colorectal cancer (CRC) patients with stages II and III CRC, which widely different in prognosis and treatment, by immunohistochemistry. PTBP1 was also upregulated in colon cancer cell lines. In our study, knockdown of PTBP1 by siRNA transfection decreased cell proliferation and invasion in vitro. Denovirus shRNA knockdown of PTBP1 inhibited colorectal cancer growth in vivo. Furthermore, PTBP1 regulates alternative splicing of many target genes involving in tumorgenesis in colon cancer cells. We confirmed that the splicing of cortactin exon 11 which was only contained in cortactin isoform-a, as a PTBP1 target. Knockdown of PTBP1 decreased the expression of cortactin isoform-a by exclusion of exon 11. Also the mRNA levels of PTBP1 and cortactin isoform-a were cooperatively expressed in colorectal cancer tissues. Knocking down cortactin isoform-a significantly decreased cell migration and invasion in colorectal cancer cells. Overexpression of cortactin isoform-a could rescue PTBP1-knockdown effect of cell motility. In summary the study revealed that PTBP1 facilitates colorectal cancer migration and invasion activities by inclusion of cortactin exon 11.

Cytoplasmic poly(A) binding protein 4 is highly expressed in human colorectal cancer and correlates with better prognosis.

Cytoplasmic poly(A) binding protein 4 (PABPC4) is an RNA-processing protein that plays an important role in the regulation of gene expression. The aim of this study was to investigate the expression pattern and identify the potential clinical significance of PABPC4 in colorectal cancer. Immunohistochemical analysis revealed that 26.7% (27/101 patients) of primary colorectal tumors and 60.5% (23/38 patients) of corresponding adjacent, normal tissues showed high cytoplasmic expression of PABPC4, whereas expression was absent in 98% (43/44 patients) of distant, normal tissues. Using Kaplan-Meier analysis, we observed that the expression of PABPC4 was significantly correlated with disease-free survival and overall survival in patients with stage II and stage III colorectal cancer (P=0.022 and P=0.020, respectively). PABPC4 expression was positively associated with survival outcome, and may have predictive value in the prognosis of patients with colorectal cancer. Taken together, our findings indicate that PABPC4 may play a role in the pathogenesis of colorectal cancer.

5-Fluorouracil and oxaliplatin modify the expression profiles of microRNAs in human colon cancer cells in vitro.

MicroRNAs (miRNAs) have recently taken center stage in the field of human molecular oncology. Most of the chemotherapeutics are able to interfere with nucleic acid metabolism and gene expression. The purpose of this study was to determine how 5-fluorouracil (5-FU) and oxaliplatin (L-OHP) modify the expression profiles of miRNAs in HCT-8 and HCT-116 colon cancer cells and whether the pharmacodynamic mechanisms of the chemotherapeutics could rely in part on their influence on miRNA expression. The expression profiles of miRNAs were determined using a miRNA microarray containing 856 human miRNA probes. The expression of selected miRNAs was then validated by real-time RT-PCR. Fifty-six up- and 50 down-regulations of miRNA expression with statistical significance were identified in colon cancer cells following exposure to 5-FU or L-OHP compared to matched control cells. The down-regulations of miR-197, miR-191, miR-92a, miR-93, miR-222 and miR-1826, whose expression was significantly down-regulated in both cell lines after the treatment of one drug or in one cell line following exposure to either drug, were further validated. Analysis of the relevant literature indicated that, in line with the tumor suppressive activity of 5-FU and L-OHP, the six down-regulated miRNAs might function as oncogenes due to their overexpression in cancers, and some of them correlated with the poor prognosis and treatment-resistance of cancer. In conclusion, we identify the modification of miRNA expression profiles in colon cancer cells following exposure to 5-FU and L-OHP, and our results indicate that their pharmacodynamic mechanisms could rely in part on their influence on the down-regulated miRNA expression. Further studies are needed to determine whether these miRNAs and their target genes might potentially provide for novel molecular markers and act as novel targets for treatment by interference.