The cytoplasmic tail of human mannosidase Man1b1 contributes to catalysis-independent quality control of misfolded alpha1-antitrypsin.

The failure of polypeptides to achieve conformational maturation following biosynthesis can result in the formation of protein aggregates capable of disrupting essential cellular functions. In the secretory pathway, misfolded asparagine (N)-linked glycoproteins are selectively sorted for endoplasmic reticulum-associated degradation (ERAD) in response to the catalytic removal of terminal alpha-linked mannose units. Remarkably, ER mannosidase I/Man1b1, the first alpha-mannosidase implicated in this conventional N-glycan-mediated process, can also contribute to ERAD in an unconventional, catalysis-independent manner. To interrogate this functional dichotomy, the intracellular fates of two naturally occurring misfolded N-glycosylated variants of human alpha1-antitrypsin (AAT), Null Hong Kong (NHK), and Z (ATZ), in Man1b1 knockout HEK293T cells were monitored in response to mutated or truncated forms of transfected Man1b1. As expected, the conventional catalytic system requires an intact active site in the Man1b1 luminal domain. In contrast, the unconventional system is under the control of an evolutionarily extended N-terminal cytoplasmic tail. Also, N-glycans attached to misfolded AAT are not required for accelerated degradation mediated by the unconventional system, further demonstrating its catalysis-independent nature. We also established that both systems accelerate the proteasomal degradation of NHK in metabolic pulse-chase labeling studies. Taken together, these results have identified the previously unrecognized regulatory capacity of the Man1b1 cytoplasmic tail and provided insight into the functional dichotomy of Man1b1 as a component in the mammalian proteostasis network.