Biology subjects

Richer, J. K.

Publications and source records attributed to Richer, J. K..

3 recordsLinked to original sources

bioRxiv2020

A single N6-methyladenosine site in lncRNA HOTAIR regulates its function in breast cancer cells

N6-methyladenosine (m6A) modification of RNA plays important roles in normal and cancer biology, but knowledge of its function on long noncoding RNAs (lncRNAs) remains limited. Here, we investigate whether m6A regulates the function of the human HOTAIR lncRNA, which contributes to multiple pro-tumor phenotypes in triple-negative breast cancer (TNBC) cells. We identify at least 8 individual m6A sites within HOTAIR, with a single site (A783) consistently methylated. Mutation of A783 impairs cellular proliferation and invasion in HOTAIR-overexpressing TNBC cells. m6A at A783 regulates HOTAIRs ability to localize to chromatin and induce gene pathways that affect tumor progression. In contrast, A783U mutant HOTAIR demonstrates loss-of-function and antimorph behaviors by impairing gene expression changes induced by WT HOTAIR and, in some cases, inducing opposite changes in gene expression. HOTAIR interacts with nuclear m6A reader YTHDC1 and high HOTAIR is significantly associated with shorter overall patient survival, particularly in the context of high YTHDC1. At the molecular level, YTHDC1-HOTAIR interactions are required for chromatin localization and regulation of gene repression. Our work demonstrates how modification of one base in a lncRNA can elicit a distinct gene regulation mechanism and drive disease-associated phenotypic changes such as proliferation and invasion.

molecular biology↗

bioRxiv2020

The capacity of the ovarian cancer tumor microenvironment to integrate inflammation signaling conveys a shorter disease-free interval

Ovarian cancer has one of the highest deaths to incidence ratios across all cancers. Initial chemotherapy is typically effective, but most patients will develop chemo-resistant disease. Mechanisms driving clinical chemo-response and -resistance in ovarian cancer are not well understood. However, achieving optimal surgical cytoreduction improves survival, and cytoreduction is improved by neoadjuvant platinum/taxane-based chemotherapy (NACT). NACT offers a window to profile pre-versus post-therapy tumor specimens, which we used to identify chemotherapy-induced changes to the tumor microenvironment. We hypothesized changes in the immune microenvironment correlate with tumor chemo-response and disease progression. We obtained matched pre- and post-NACT archival tumor tissues from patients with high-grade serous ovarian cancer (patient n=6). We measured mRNA levels of 770 genes (NanoString), and performed reverse phase protein array (RPPA) on a subset of matched tumors. We examined cytokine levels in additional pre-NACT ascites samples (n=39) by multiplex ELISA. A tissue microarray with 128 annotated ovarian tumors expanded the transcriptional, RPPA, and cytokine data by multi-spectral immunohistochemistry. In NanoString analyses, transcriptional profiles segregated based on pre- and post-NACT status. The most upregulated gene post-NACT was IL6 (17.1-fold, adjusted p = 0.045). RPPA data were highly concordant with mRNA, consistent with elevated immune infiltration. Elevated IL-6 in pre-NACT ascites specimens correlated with a shorter time to recurrence. Integrating NanoString, RPPA, and cytokine studies identified an activated inflammatory signaling network and induced IL6 and IER3 (Immediate Early Response 3) post-NACT, associated with poor chemo-response and decreased time to recurrence. Taken together, multi-omic profiling of ovarian tumor samples before and after NACT provides unique insight into chemo-induced changes to the tumor and microenvironment. We integrated transcriptional, proteomic, and cytokine data and identified a novel IL-6/IER3 signaling axis through increased inflammatory signaling which may drive ovarian cancer chemo-resistance.

cancer biology↗

bioRxiv2020

Estrogen receptor alpha mutations in breast cancer cells cause gene expression changes through constant activity and through secondary effects

While breast cancer patients with tumors that express estrogen receptor (ER) generally respond well to hormone therapies that block ER activity, a significant number of patients relapse. Approximately 30% of these recurrences harbor activating mutations in the ligand binding domain (LBD) of ER, which have been shown to confer ligand-independent function. However, much is still unclear regarding the effect of mutant ER beyond its estrogen independence. To investigate the molecular effects of mutant ER, we developed multiple isogenic ER mutant cell lines for the most common LBD mutations, Y537S and D538G. These mutations induced differential expression of thousands of genes, the majority of which were mutant allele-specific and were not observed upon estrogen treatment of wildtype cells. These mutant-specific genes showed consistent differential expression across ER mutant lines developed in other laboratories. Wildtype cells with long-term estrogen exposure only exhibited some of these transcriptional changes, suggesting that mutant ER causes novel regulatory effects that are not simply due to constant activity. While ER mutations exhibited minor effects on ER genomic binding, with the exception of ligand independence, ER mutations conferred substantial differences in chromatin accessibility. Mutant ER was bound to approximately a quarter of mutant-enriched accessible regions that were enriched for other DNA binding factors including FOXA1, CTCF, and OCT1. Overall, our findings indicate that mutant ER causes several consistent effects on gene expression, both indirectly and through constant activity.

cancer biology↗