Grant Program Winners for miRNA functional studies
SwitchGear Genomics is pleased to announce our grantees for the SwitchGear Grant Program for miRNA functional studies. Meet our winners below!
Who: Dr. Gerardo Ferbeyre
Institution: University of Montreal
Product Highlight: miR-122, miR-155, miR-21, miR-29a, miR-124, miR-200a, miR-223, miR-208a, let-7a GoClone miRNA target sets
We are interested in creating new miRNAs. These miRNAs are designed by our computer program MultiTar. Using MultiTar we can choose the targets for our artificial miRNAs. In this way we can create reagents to inhibit gene combinations that are not altered by any single known miRNA (NAR in press).
We will use 3’UTR GoClones to validate the functionality of our artificial miRNAs (called smart RNAs). We request five reporters, which are targets of five different natural miRNAs. Then we will use MultiTar to design several smart RNAs against these five reporters. We will construct retroviral expression vectors for the five smart RNAs, all potentially capable of recognizing the five 3’UTRs selected form the Switchgear collection. The expression vectors will be introduce in 293 cells by retroviral gene transfer. The expression level of each smart RNA will be measured by qPCR and their functionality will be evaluated by transfecting each 3’UTR goclones into the cell line expressing each of the five smart RNAs. We expect to learn more about miRNA targeting from the results of these experiments and apply the new knowledge to further improve MultiTar.
Who: Dr. Ying Zhang
Institution: University of Massachusetts Medical School
Product Highlight: 3′UTR GoClone reporters
In my studies, I characterize miRNAs that regulate skeletal development by expression profiling of miRNAs in differentiating osteoblasts. Predictions of target genes for prominently regulated miRNAs are being validated for robust inhibition of target gene expression at the protein and/or mRNA levels. However, it is necessary to examine miRNA effects in target gene reporters containing full length 3’UTRs to establish which genes are true direct targets of specific miRNAs. The gene-specific 3’UTR GoClone reporters and control constructs available from SwitchDB are critical for studies on miRNA dependent control of target gene expression during skeletogenesis.
Dr. Ying Zhang is a postdoctoral follow in Department of cell biology at University of Massachusetts Medical School. Her studies focus on transcriptional regulation of bone-related genes and microRNAs regulation of bone formation.
Who: Dr. Leon van Kempen
Institution: Radboud University Nijmegen Medical Center, Dept. of Pathology
Product Highlight: miR-122 GoClone target set
Loss of E-cadherin expression in melanoma correlates with aggressiveness and poor survival. Our research group aims to understand the mechanism by which melanoma cells gain their highly metastatic properties. We and others have demonstrated that miR200a increases the expression of the E-cadherin transcriptional repressor ZEB1/2 and that this strongly correlates with poor prognosis of melanoma. TGFb1 is a modulator of miR200a expression, but the molecular underpinning that drives miR200a expression remains largely elusive. We have generated cell lines that are responsive to TGFb1 and in which we can modulate the expression of TGFb1-reponsive transcription factors. Using the GoClone assay, we can now analyze the activity of miR200a in the presence or absence of steady state or TGFb1-activated transcription factor levels. This will yield important information about the transcription factors that modulate miR200a expression.
Who: Dr. Li Jia
Institution: Washington University in St. Louis
Product Highlight: 3′UTR GoClone reporters
Aberrant expression of microRNA 221/222 has been investigated in many cancers. We recently observed over-expression of miR221/222 in androgen-independent prostate cancer (PCa) cells, which indicated their important roles in PCa progression. The potential target genes for miR221/222 include CDKN1A, CDKN1B, and CDKN1C. In this application, we intend to examine miR221/222 activities using 3′UTR luciferase reporters to in different PCa cell lines.
(1) Five PCa lines will be used: RWPE, LAPC4, LNCaP, C4-2B, and 22Rv1 cells represent from normal prostate epithelial cells to androgen-dependent and –independent PCa cells, which mimic clinical progression.
(2) We will test four 3′UTR target reporters (CDKN1A, CDKN1B, CDKN1C, and AR) along with 2 controls (GAPDH and empty vector) in a transient transfection system using luciferase assays.
(3) We will also test miR221/222 activities in the absence/presence of dihydrotestosterone (DHT) to further study androgen-mediated regulatory effect on miR221/222 target genes.