Talk abstracts
Talk on Saturday 09:30-09:45am submitted by Pavithra Madamarandawala
Parkinson's Disease associated genes are directly regulated by Pumilio proteins
Pavithra S. Madamarandawala (Department of Biology, University of Missouri-St. Louis), Wendy M. Olivas (Department of Biology, University of Missouri-St. Louis)
Abstract:
Parkinson’s Disease (PD) is caused by the loss of dopamine producing neurons and is the second most common neurodegenerative disorder. In addition to well-known PD-associated mutations, overexpression of several genes is linked to onset and severity of PD, including SNCA, encoding alpha-synuclein that aggregates during PD, and LRRK2, a kinase associated with mitochondrial dysfunction. The Pumilio (PUM) family of eukaryotic RNA-binding proteins is involved in post-transcriptional regulation of mRNA. PUMs bind to a conserved region called a PUM/Puf recognition element (PRE), which is commonly present in the 3’ untranslated regions (UTRs) of target mRNAs. The canonical human PRE is UGUA(A/U)AUA, but binding can also occur at some non-canonical sites. Upon binding, PUM recruits mRNA decay machinery to the mRNA and inhibits translation. The 3’UTRs of LRRK2 and SNCA each have at least one canonical PRE, and our preliminary work demonstrated that knockdown of the two PUM proteins (PUM1, PUM2) in the human neuroblastoma cell line, SH-SY5Y, results in overexpression of LRRK2 and SNCA. To elucidate this regulation, we conducted luciferase assays using reporters under the control of the 3’UTR of SNCA or LRRK2. Luciferase expression of each reporter was elevated in response to PUM1/2 KD, confirming that PUM1/2 regulation is directly acting through the 3’UTRs. Further, using mutant luciferase reporters (changing the UGUA to ACAC), we found that both the canonical and non-canonical PREs of SNCA, and the three canonical PREs of LRRK2 are all involved in PUM1/2-mediated regulation. A further increase in expression in all double PRE mutant combinations demonstrated an additive effect of regulation by each PRE. Our current work is analyzing combinatorial regulation of the SNCA and LRRK2 mRNAs by PUMs and miRNAs. Overall, these results demonstrate the direct role of PUM1/2 in regulation of PD-associated genes and paves the way to further understand additional regulatory mechanisms.
Keywords: Pumilio, RNA decay, RNA binding protein