Protein Quality Control in an Aging Disorder

Human aging, a time-dependent progressive decline in the function of cells and tissues, has intrigued scientists throughout history. This extremely complex, multifactorial process is the biggest risk factor for many diseases such as cancer as well as cardiovascular and neurodegenerative diseases. Since several human premature aging-like syndromes are characterized by features resembling normal aging, they provide important insights into the molecular mechanisms underlying human aging. The most pronounced premature aging disorder is Hutchinson-Gilford Progeria Syndrome (HGPS), an extremely rare, autosomal dominant genetic disorder reflecting several aspects of normal aging, such as loss of hair and subcutaneous fat, aged-looking skin, joint stiffness, as well as severe osteoporosis, atherosclerosis and cardiovascular diseases. Classical HGPS is caused by a de novo heterozygous mutation (1824C>T, p.G608G) in exon 11 of LMNA, the gene encoding A-type lamins. This mutation activates a cryptic exonic splice site resulting in the expression of a mutant form of lamin A termed progerin. Progerin acts in a dominant-negative fashion and induces numerous cellular defects, including highly lobulated nuclei with thickened lamina, changes in chromatin organization and gene expression, compromised cell-cycle regulation and premature senescence. The molecular mechanisms leading to the cellular defects in HGPS are unknown. In this proposal, I hypothesize that progerin exerts its deleterious function through accumulation and aggregation in the cell nucleus, which in turn affects protein homeostasis and in this way contributes to disease symptoms. I will test the effect of progerin on two main mechanisms involved in the maintenance of the proteome: 1. Cellular chaperones, which are essential for proper folding of nascent proteins and for preventing protein aggregation, and 2. the proteasomal system which removes damaged and misfolded proteins. Specifically, I will ask whether recruitment of chaperones to the progerin aggregates results in impaired folding of nascent proteins elsewhere in the cell and to a decreased ability of the cell to dissociate newly formed aggregates. I will also test whether mis-regulation in the chaperone network contributes to the increased progerin accumulation in the nucleus. Furthermore, I will probe whether increased progerin accumulation can inhibit the activity of the ubiquitin-proteasome system (UPS) by saturating the capacity of one or more chaperones required for proper UPS function, or by direct interaction with the proteasome. In parallel, I will test whether progerin is targeted for proteasomal degradation, the main pathway involved in protein clearance from the cells. These lines of investigation address a timely and important question in the field and will yield new insights into the pathology of the disease. In the long term, they have the potential to provide the basis for the development of new treatments for HGPS.

Hutchinson-Gilford Progeria Syndrome (HGPS) is an extremely rare premature aging disorder reflecting several aspects of normal aging, such as loss of hair and subcutaneous fat, aged- looking skin, joint stiffness, as well as severe osteoporosis, atherosclerosis and cardiovascular diseases. Classical HGPS is caused by constitutive production of progerin, a mutant form of the nuclear architectural protein lamin A. Progerin causes numerous cellular defects, including highly lobulated nuclei with thickened lamina, changes in chromatin organization and gene expression, compromised cell-cycle regulation and premature senescence. The molecular mechanisms leading to the cellular defects in HGPS are still poorly understood. The goal of my work was to understand the effect of progerin accumulation on protein quality control and how the impairment of this pathway contributes to the progression of the cellular HGPS phenotypes. I hypothesized that progerin exerts its deleterious function, in part, through its accumulation and aggregation in the nucleus and proposed a mechanism by which progerin-containing protein aggregates recruit and tether molecular chaperones, affecting protein homeostasis in the rest of the cell. My data indicate that the Hsp110 and Hsc70 cytosolic chaperones accumulate at the endoplasmic reticulum (ER) of progerin-expressing cells. This re-localization is accompanied by the activation of an adaptive response to ER stress, including an increase in various ER chaperones, transcriptional activation of Unfolded Protein Response (UPR) genes and increased phosphorylation of IRE1a and PERK, two major mediators of the UPR pathway. In addition, immunofluorescence analysis and super resolution microscopy reveal localization of several ER chaperones at the nuclear periphery where they co-localize with progerin. The ER chaperone GRP78/BiP co-immunoprecipitates with GFP-progerin but not with the wild type lamin A, suggesting that progerin interacts, directly or indirectly, with the resident ER chaperone GRP78/BiP. Prolonged ER stress coupled with an inadequate UPR response has been linked to apoptosis of vascular smooth muscle cells, atherosclerosis and cardiovascular diseases, phenotypes commonly observed in HGPS patients. In line with the observed upregulation of the UPR pathway in cultured cells, a significant upregulation of several UPR genes was observed in the heart and aorta of progeria mice. In summary, my findings shed new light on the molecular mechanisms that govern HGPS pathology and may provide a basis for the development of new treatment strategies for this still incurable disease. In the long term, understanding mechanisms underlying rapid aging disorders could not only contribute to our basic knowledge of various processes in health and disease, but also to a better understanding of the aging process itself.