Analysis of the transcriptional activation and post‐transcriptional regulation of transient receptor potential mucolipin‐2 gene (561.5)

Abstract

Mucolipidosis type IV (MLIV) is a neurodegenerative disease caused by loss or dysfunction of the human transient receptor potential mucolipin 1 (TRPML1) protein. TRPML1 is a non‐selective cation channel related to two other proteins, TRPML2 and TRPML3. The high degree of sequence and functional similarities between TRPML1 and TRPML2 make TRPML2 an excellent candidate to potentially substitute for the loss of TRPML1 function in MLIV. In order to consider this possibility, we first needed to study the mechanisms that control the expression of TRPML2 gene. We cloned DNA regions of varying size upstream of the TRPML2 gene transcriptional start site (TSS) into a dual luciferase reporter assay. We identified a putative core promoter region between ‐320 and ‐287 of the TSS containing a consensus sequence for six transcription factors. Heterologous expression of the candidate TFs conferred an effect on the expression of endogenous TRPML2 transcripts compared to control. Meanwhile, we found that a specific micro‐RNA was responsible for post‐transcriptional regulation of endogenous TRPML2 transcripts as evidenced by real‐time quantitative PCR (qPCR) technique. We then screened a drug library and confirmed by real‐time qPCR that each drug candidate significantly increased the expression of endogenous TRPML2 transcripts. These results show that we could induce the expression of the TRPML2 gene through exogenous manipulation of transcriptional activators or post‐transcriptional regulators. Understanding the mechanism of TRPML2 expression is a step closer to our goal of using TRPML2 protein to substitute for the loss of TRPML1 protein function in MLIV.Grant Funding Source: Supported by NIH R15‐NS070774 and CSUF Intramural Grants