Posttranslational modifications of serine protease TMPRSS13 regulate zymogen activation, proteolytic activity, and cell surface localization

Abstract

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Impact of SARS-CoV-2 Spike Mutations on Its Activation by TMPRSS2 and the Alternative TMPRSS13 Protease

“…TMPRSS2 and TMPRSS13 showed a strikingly similar staining pattern, with weak staining of the plasma membrane and apparently high abundance in the endoplasmic reticulum (ER) and secretory pathway. The same picture was seen by another group, who confirmed the ER localization of expressed TMPRSS13 by costaining with an anti-ER antibody (31). We observed no plasma membrane staining under nonpermeabilized conditions, in full accordance with another study using a C-terminus (in this case V5 tag)-directed antibody (32).…”

“…In this context, we refer to the link between spike fusogenicity and SARS-CoV-2 virulence, observed in animal studies with the Delta and Omicron variants (5,7,8). Second, we are puzzled by the observation that the TMPRSS2 and TMPRSS13 proteins expressed in HEK293T cells gave relatively weak staining of the plasma membrane but appeared to localize mainly in the ER and secretory pathway (as already seen by others [31,45]). Since both undergo (auto)cleavage and shedding of the extracellular protease domain, it remains to be determined in which form these proteases conduct S29 cleavage during virus entry.…”

“…The band of ~55 kDa represents full-length TMPRSS2 ( 30 ), while that of ~32 kDa represents the serine protease domain resulting from autocleavage. For TMPRSS13, we observed two dominant bands: full-length TMPRSS13 having, in glycosylated form, a molecular weight (MW) of ~70 kDa and the presumed intracellular fragment of ~26 kDa that results from autocleavage ( 31 ).…”

Transmembrane serine protease TMPRSS2 implicated in SARS-CoV-2 infection is autoactivated intracellularly and requires N-glycosylation for regulation

“…N-glycosylation is an important mechanism in the regulation of TTSP folding, intracellular trafficking, and zymogen activation ( 41 , 52 , 53 ). However, the functionality of individual N-glycans in a particular protein is hard to predict.…”

“…Most TTSPs are synthesized in a single-chain zymogen form. Posttranslational modifications, including N- or O-glycosylation, zymogen activation, phosphorylation, and ectodomain cleavage, are important mechanisms in regulating TTSP activities ( 37 , 38 , 39 , 40 , 41 , 42 , 43 , 44 , 45 ). Previous studies indicate that TMPRSS2 is synthesized as a single-chain zymogen and activated by autocatalysis ( 46 ).…”