This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.Transcript levels for various enzymes and proteins involved in human glycoprotein biosynthesis, ER-associated glycoprotein disposal (ERAD), and unfolded protein response (UPR) will be quantitated by qRT-PCR. A set of projects are being initiated in collaboration with Dr. Hung Do (Amicus Therapeutics) to examine the roles of individual genes in ER quality control and glycoprotein degradation in human cells. Amicus Therapeutics is a biotech company developing small molecule chemical chaperone therapeutics for treatment of lysosomal storage diseases. These diseases are characterized by the accumulation of misfolded lysosomal enzymes in the ER of human cells where they potentially induce an unfolded protein response. We will employ patient-derived lysosomal storage disease cell lines as a model system to examine the effects of treatment with chemical chaperones or perturbation of the machinery involved in ER-associated degradation (by over-expression or shRNA knockdown). The effects of these treatments on lysosomal enzyme folding, maturation, and transport versus enzyme degradation will be examined in one set of studies while another set of studies will examine the effects of these treatments on transcript levels of the full set of glycan-related genes as well as genes involved in ERAD and UPR. These studies should reveal the interplay between misfolded glycoprotein biosynthesis, cellular glycosylation, ER degradation machinery, and machinery of the unfolded protein response.
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