Date of Award
Doctor of Philosophy (PhD)
Adam D. Linstedt
Shiga toxin (STx)–producing bacteria including Shigella dysenteria and the shigatoxigenic group of Escherichia coli (STEC) cause severe diarrhea that can develop into life-threatening hemolytic uremic syndrome. STx invades cells by binding to a cell surface receptor and undergoing endocytosis into early endosomes. Instead of subsequent lysosomal degradation, it actively sorts into early endosomal tubules by binding GPP130 (Golgi Phosphoprotein of 130KDa) a cis- Golgi resident integral membrane protein that constitutively cycles between early endosomes and Golgi. This directs the toxin to the Golgi where it continues its retrograde trafficking to exert its cytotoxic effects. Interestingly, elevated, non-toxic levels of manganese (Mn) protect cells from STx induced death via downregulation of GPP130. Mn binds to GPP130 in the Golgi, causes GPP130 to oligomerize/aggregate and the oligomerized protein gets diverted to lysosomes. Surprisingly, based on experiments using the self-interacting FM domain, it appears generally true that oligomerization of a Golgi protein can switch its trafficking itinerary leading to its lysosomal degradation via a GGA1-clathrin dependent pathway. However, how this sorting switch at the Golgi is mediated remains unknown. This has important implications for our understanding of regulation of protein trafficking as well as quality control pertaining to aggregated proteins in the Golgi. Here we provide evidence that Mn-induced exit of GPP130 from the Golgi towards lysosomes is mediated by the sorting receptor, sortilin. Sortilin was found to interact with GPP130 in cells within a ~50 residue stretch of the GPP130 lumenal stem domain. Interestingly, Mn induced turnover of another Golgi membrane protein, TMEM165 was also found to be sortilin-dependent. In contrast, FM-induced lysosomal trafficking of GPP130 as well as the Golgi protein galactosyltransferase (GT) was sortilin independent and occurred even in the absence of the native lumenal domain of GT. Thus, sortilin-dependent as well as sortilin-independent sorting mechanisms target aggregated Golgi membrane proteins for lysosomal degradation.
Venkat, Swati, "Sorting of Oligomerized Proteins: Implications in Toxin Trafficking and Quality Control" (2017). Dissertations. 1077.