The University of Texas MD Anderson Cancer Center UTHealth Graduate School of Biomedical Sciences Dissertations and Theses (Open Access)
THE ROLE OF PHOSPHORYLATION IN PAM2 MOTIF-CONTAINING PROTEINS MEDIATED MESSENGER RNA DEADENYLATION
Date of Graduation
Biochemistry and Molecular Biology
Doctor of Philosophy (PhD)
Ann-Bin Shyu, Ph.D.
Zheng Chen, Ph.D.
Jianping Jin, Ph.D.
William W. Mattox, Ph.D.
Ambro Van Hoof, Ph.D.
Phosphorylation regulates many cellular processes. However, its role in mRNA deadenylation, a process to remove poly adenosines from the mature mRNA 3’ end tail, is unclear. The length of poly(A) tail determines mRNA stability and translation efficiency. Poly(A)-binding protein (PABP), which binds to newly synthesized poly(A) tails homogeneously and is known as a scaffold protein for PAM2 motif-containing proteins, plays a pivotal role in the shortening of poly (A) tails. This study is to examine the role of phosphorylation of PAM2 motif–containing proteins in regulating their interactions with PABP and mRNA deadenylation function.
The PAM2 motif, a region required for binding PABP C-terminal domain, is embedded inside the intrinsically disordered region (IDR) of its containing proteins. IDR is known to be a favored site for phosphorylation, and the PAM2 motif is surrounded by numerous potential phosphorylation sites. To test whether phosphorylation plays a role in controlling the interaction between PAM2-containing proteins and PABP, this study used complementary approaches (to alter the phosphorylation level of PAM2 motif-containing proteins biochemically and to create phospho-mimetic (PM) and non-phosphorylatable (NP) mutants). The results showed that reducing or increasing phosphorylation of these proteins can enhance or diminish their interaction with PABP, respectively. Furthermore, diminished PABP interaction compromises the biological functions of these proteins in deadenylation and miRNA-mediated gene silencing.
Using Tob2-PABP interaction as a model, this study provides more insight into the mechanism by which phosphorylation of PAM2 motif–containing proteins regulates their interactions with PABP. The data presented here suggest that the two PAM2 motifs of Tob2 interact with PABP in a synergistic rather than addictive manner. Multiple phosphorylation induced by JNK1 in Tob2 IDR compromises Tob2-PABP interaction. A critical site for phosphorylation of Tob2 was identified. Global mRNA deadenylation can be regulated by phosphorylation status of this site.
Collectively, this work provides a new concept, in which signaling pathways crosstalk may tune global deadenylation rates through phosphorylation of PAM2 motif-containing proteins to modify their PABP affinity.
mRNA deadenylation, PAM2 motif, Phosphorylation, Intrinsically disordered regions, PABP, Tob2, Pan3, Tnrc6c, JNK, CDK