Haploinsufficiency of protein-coding genes as an age-related disease mechanism

2013 new Scholar Award in aging

Recent advances in technologies for determining DNA sequences have revealed that every human genome contains small deletions, as well as mutations that abolish the function of one of two copies of tens of genes. Such mutations and deletions do not result in complete loss of function, but rather in reduced dosage of the protein products of the affected genes. The effects of reduced gene dosage (referred to as gene haploinsufficiency) on the cell or organism remain to date poorly studied. Evidence suggests that haploinsufficiency may produce phenotypes solely or predominantly in the presence of additional “stressors”. We propose that one such “stressor” is the decline of protein turnover (homeostasis) induced by aging. In the proposed research we will test the hypothesis that an aging-related decline of protein activity decreases the tolerance to the haploinsufficiency of genes that code for proteins and results in dysregulation of cellular functions that are well-compensated for earlier in life. This can render phenotypes of gene haploinsufficiency at the cellular and organismal level conditional upon aging. Genes whose function is dependent on dosage can only be revealed by functional assays. Our laboratory harnesses breakthrough stem cell technologies, namely somatic cell reprogramming and genetic engineering, to develop new models of normal and abnormal hematopoiesis that enable functional genetic studies, hitherto unthinkable for the human genome. We are deriving induced pluripotent stem cells (iPSCs) – cells that are practically identical to pluripotent stem cells (PSCs) found in early embryos – directly from cells of adult patients. We are also engineering genetic and chromosomal abnormalities that are believed to cause or predispose to diseases in normal PSCs. We are then using step-wise protocols (directed differentiation) to derive blood cells from these PSCs and assess their phenotype. Using these technologies we have recently developed a novel model of an age-related disorder of the hematopoietic system, myelodysplasia (MDS). Over 87% of MDS cases arise after the age of 60 years. There is strong evidence that deletion of part of one copy of chromosome 7 plays a fundamental role in this disease. We have therefore derived iPSCs from patients with MDS and genetically engineered iPSCs with chromosome 7 deletions. This model provides a novel platform to study basic mechanisms of aging and offers unprecedented opportunities for functional genetics studies in human cells. Here we will use it to seek evidence for an increased dependence of protein function on gene dosage upon aging. Specifically, we will characterize the cellular phenotypes conferred by loss of one copy of chromosome 7 in our MDS-PSC model and identify haploinsufficient genes residing on the long arm of chromosome 7 that mediate those phenotypes. We will then test the effects of aging in the ability of blood cells to compensate for haploinsufficiency of these genes, by reducing (knocking down) their expression in stem cells of the blood of healthy individuals of different age groups. Our study may implicate haploinsufficiency of protein-coding genes as a general mechanism of aging of the hematopoietic system and of aging-associated disorders more generally and provide new insights into the genetics of aging and healthspan. It may point to new opportunities for therapeutic interventions aimed at restoring the levels and/or function of protein products of haploinsufficient genes that are critical for aging and aging-related diseases and lead to further studies exploring the impact of environmental factors in modifying their levels and/or function.

Researchers
Eirini Papapetrou Ph.D.
Washington, University of