Poster abstracts
Poster number 173 submitted by Bo Zhang
Identification of the mechanism for PABPN1 translational repression during cardiac development and hypertrophy
Bo Zhang (Department of Biochemistry, University of Illinois at Urbana-Champaign), Joseph Seimetz, Chaitali Misra, Emelia Smith (Department of Biochemistry, University of Illinois at Urbana-Champaign), Qinyu Hao, Kannanganattu V. Prasanth (Department of Biochemistry, University of Illinois at Urbana-Champaign), Auinash Kalsotra (Department of Biochemistry, University of Illinois at Urbana-Champaign)
Abstract:
Cardiomyocytes are the muscle cells that make up the heart muscle. These contractile cells occupy ~70-85% the volume of the mammalian heart and enable the heart to pump blood throughout the body. The majority of cardiomyocytes become post-mitotic soon after birth and since then the heart increases in size through the expansion of the volume of existing cardiomyocytes, namely postnatal hypertrophy or maturational hypertrophy. As the animal reaches adulthood, cardiomyocytes stopped expansion and matured; at the same time, protein synthesis in these cells dramatically decreases. However, upon particular stimuli, protein synthesis surges in the mature cardiomyocytes, accompanied with physiological or pathological hypertrophy of these cells and changed heart physiology. Recently, our lab has identified the poly(A)-binding protein nuclear 1 (PABPN1) as an important regulator of cardiac hypertrophy. PABPN1 is responsible for the elongation of poly(A) tail and is involved in multiple aspects of mRNA metabolism including nucleocytoplasmic trafficking and nonsense-mediated decay (NMD). The cardiac PABPN1 levels dramatically dropped upon neonatal to adult transition. Elevated expression of PABPN1 was not only observed in mouse cardiac hypertrophy models, but also found in failing human hearts, suggesting a tight connection between postnatal re-expression of PABPN1 and pathological cardiac hypertrophy. Forced expression of this protein in adult cardiomyocytes induced severe cardiac hypertrophy, impaired contractile force and sudden death of the experimental animals. Interestingly, despite of the dramatic reduction of PABPN1 protein in the adult animals, the RNA level of Pabpn1 was hardly changed. However, little is known about the underlined mechanism for this translational regulation of Pabpn1 expression.
We hypothesize that nuclear sequestration caused by insufficient splicing of intron results into translational arrest of PABPN1 in mature striated muscle cells. We found that the repression of PABPN1 translation was achieved by the sequestration of Pabpn1 mRNA in the nucleus. RNA fluorescent in situ hybridyzation (RNA-FISH) and nucleus-cytoplasm fractionation suggested that Pabpn1 transcripts were preferably localized to the nucleus in the adult cardiomyocytes and differentiated myotubes. We have identified the intron 6 of Pabpn1 as a potential cis-acting element that is responsible for the translational regulation. We observed increased intron 6 detention in adult heart and differentiated myotubes. These intron 6-detained transcripts preferably localized to the nucleus. We have also identified the splicing factor Rbfox2 as a potential trans-acting factor for this regulation. A crosslinking and immunoprecipitation (CLIP) assay revealed that RBFOX2 bound to the intron 6 regain of Pabpn1 transcript. Interestingly, both PABPN1 protein levels and Pabpn1 mRNA splicing increased when RBFOX2 was overexpressed. Currently, we are in the process of using non-biased methods like RNA immunoprecipitation (RIP) and RNA antisense puarification-mass spectrometry (RAP-MS) RAP to verify this finding and to further exploit other potential candidates for this regulation. The identification of the mechanism for hypertrophic growth of cardiomyocyte is important for our understanding of cardiac development and hypertrophic pathology and potentially will benefit the prevention and treatment of heart disease.
Keywords: PABPB1, translational repression, cardiac development and hypertrophy