The ability to strictly regulate apoptosis is particularly important for postmitotic cells such as neurons, cardiomyocytes and myotubes because these cells have limited regenerative potential and are maintained for the lifetime of the organism. Failure to restrict apoptosis can result in increased vulnerability to cell death, as seen in many degenerative diseases. Therefore, an understanding of survival mechanisms in postmitotic cells is of physiological and pathological importance. The mitochondrial release of cytochrome c (cyt c) is a crucial event that triggers caspase activation during apoptosis. Here, we have identified a novel mechanism for regulating cyt c-mediated apoptosis engaged by neurons and other postmitotic cells, where cytosolic cyt c itself is targeted for proteasomal degradation. Importantly, while we do not observe cyt c degradation in primary mitotic cells such as fibroblasts, we find mitochondrial-released cyt c to be also degraded in certain cancer cells. As evasion of apoptosis is an important feature of both postmitotic and cancer cells, targeting cytosolic cyt c for degradation could be a shared mechanism used by these cells for survival. In this proposal, our goals are to identify the molecular mechanisms by which cytosolic cyt c is targeted for degradation.
In Aim 1, we will define the factors which determine whether or not a cell will target cytosolic cyt c for degradation, including examining whether low levels of Apaf-1 in cells is a key determinant for cyt c degradation.
In Aim 2, we will examine cyt c ubiquitination and identify the specific residues of cyt c that are targeted for ubiquitination. Our hypothesis is that ubiquitinated cyt c is not able to bind Apaf-1. We will test this, and examine whether conditions that block cyt c degradation increase the vulnerability of postmitotic cells to apoptosis. Our focus in Aim 3 is to identify the E3 ubiquitin ligase that targets cyt c for degradation. Our preliminary results suggest that cyt c binds to Hsp70 and the E3 Ligase CHIP (Carboxyl Heat shock protein 70-Interacting Protein) in the cytosol. Therefore, we will test whether the ubiquitination and degradation of cytosolic cyt c is mediated by CHIP in postmitotic and cancer cells.

Public Health Relevance

Our plans here are to investigate how the cell death pathway is regulated in mammalian cells. We have discovered a novel mechanism engaged by postmitotic cells such as neurons, cardiomyocytes and myotubes that highly restricts cell death and likely ensures their long term survival. Understanding the survival mechanisms used by postmitotic cells has enormous clinical significance because increased death of these cells is central to the pathology of many neurodegenerative diseases, cardiac pathologies and muscular dystrophies.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM078366-07
Application #
8323354
Study Section
Special Emphasis Panel (ZRG1-BDA-A (02))
Program Officer
Maas, Stefan
Project Start
2006-09-01
Project End
2014-08-31
Budget Start
2012-09-01
Budget End
2013-08-31
Support Year
7
Fiscal Year
2012
Total Cost
$305,022
Indirect Cost
$97,122
Name
University of North Carolina Chapel Hill
Department
Physiology
Type
Schools of Medicine
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Gama, Vivian; Swahari, Vijay; Schafer, Johanna et al. (2014) The E3 ligase PARC mediates the degradation of cytosolic cytochrome c to promote survival in neurons and cancer cells. Sci Signal 7:ra67
Gama, Vivian; Deshmukh, Mohanish (2013) Adenosine: essential for life but licensed to kill. Mol Cell 50:307-8
Crowther, Andrew J; Gama, Vivian; Bevilacqua, Ariana et al. (2013) Tonic activation of Bax primes neural progenitors for rapid apoptosis through a mechanism preserved in medulloblastoma. J Neurosci 33:18098-108
Garcia, I; Crowther, A J; Gama, V et al. (2013) Bax deficiency prolongs cerebellar neurogenesis, accelerates medulloblastoma formation and paradoxically increases both malignancy and differentiation. Oncogene 32:2304-14
Smith, Michelle I; Huang, Yolanda Y; Deshmukh, Mohanish (2009) Skeletal muscle differentiation evokes endogenous XIAP to restrict the apoptotic pathway. PLoS One 4:e5097
Vaughn, Allyson E; Deshmukh, Mohanish (2008) Glucose metabolism inhibits apoptosis in neurons and cancer cells by redox inactivation of cytochrome c. Nat Cell Biol 10:1477-83