Rabbit Anti-Human MYC (Phospho-Ser373) polyclonal antibody

Reproductive hormone imbalance in infertile women is correlated to high levels of phthalates and alkylphenols, which are among endocrine-disrupting chemicals (EDCs). Previous studies have shown that they interfere with gene expression by deregulating levels of microRNAs (miRs), small non-coding RNAs targeting mRNAs encoding enzymes in the hormone biosynthesis pathway. However, this effect depends on the target organ, dose and whether or not they are alone or in mixtures. Our goal was to study whether the biosynthesis, and a specific group of miRs targeting mRNAs encoding enzymes in steroid hormone biosynthesis, are deregulated in the ovaries of female mice chronically exposed to a mixture of three phthalates (DEHP+DBP+BBP) and two alkylphenols (NP+OP) at a human environmentally relevant dose. We performed qPCR and Western blot assays along with a bioinformatics approach and found that this mixture modified the biogenesis machinery of miRs, inducing an increase in the mRNA levels of Drosha and Dicerl and DROSHA protein levels. In addition, we found changes in the precursor and mature forms of miR-96-5p, miR-200b-3p, miR-365-3p, miR-378a-3p and miR-503-5p which target steroidogenic pathway enzymes. Finally, using primary granulosa cell culture, we confirmed that miR-200b-3p targets Cyp19a1, transcript encoding CYP19A1, the enzyme that produces estradiol (E-2). These results indicate that chronic exposure to phthalates and alkylphenols mixture alters the biogenesis of ovary miRs and increases the expression of miRs implicated in the control of steroidal hormone synthesis in female mice, thus contributing to reproductive pathologies.

Epidermal growth factor (EGF)-receptor regulates several downstream signaling pathways upon EGF stimulation that involves cell proliferation, migration and invasion. Internalized EGF-receptor is either recycled or degraded, which fate is regulated in part by Ras interference 1 (RIN1). In this study, we tested the hypothesis that RIN1, a Ras effector protein and Rab5 guanine nucleotide exchange factor, controls several signaling molecules leading to the modulation of the telomerase activity; thus, allowing proper cell proliferation. We report that expression of RIN1 completely blocked proliferation of MCF-12 A and MCF-7 cells, while partially inhibited proliferation of MDA-MB-231 cells upon EGF stimulation. Furthermore, expression of the C-terminal region of RIN1 selectively plays a critical role in the inhibition of the proliferation of MDA-MB-231 cells. However, this inhibitory effect was specifically affected by the independent expression of RIN1:Vsp9 and RINLRA domains. Additionally, endogenous level of expression of RIN1 was decreased in metastatic MDA-MB-231 cells as compared with nontumorigenic MCF-12 A cells. We observed that expression of RIN1:R94A mutant blocked the proliferation of MDA-MB-231 cells, while expression of RIN1:Y561F and RIN1:R629A mutants completely reversed the inhibitory effect of RIN1:WT. Consistent with our observations, we found that expression of RIN1:WT in MDA-MB-231 cells diminished both protein kinase B (AKT) and extracellular-signal-regulated kinase 1/2 (ERK1/2) activities while p38 mitogen-activated protein kinases (p38 MAPK) and stress-activated protein kinase (SAPK)/c-Jun N-terminal kinase (JNK) were unaffected, but it produced downregulation of cellular-myelocytomatosis (c-Myc), erythroblast transformation specific (Ets2) and signal transducer and activator of transcription 3 (Stat3) activities. Inversely, expression of high-mobility group box 1 (HMBG1) was inhibited whereas expression of forkhead box transcription factor 1 (FOXO1) was increased in cells expressing RIN1. Interestingly, expression of RIN1 blocked telomerase activity and human telomerase reverse transcriptase (hTERT) expression, which correlated with the downregulations of c-Myc, Ets-2 and Stat3 activation. Taken together these findings indicate that RIN1 is a critical player in the modulation of the telomerase activity as well as hTERT expression in MDA-MB-231 cells upon EGF stimulation.