Analysis of the polycystin complex (PCC) in human urinary exosome–like vesicles (ELVs)

Wendy A. Lea, Kerri McGreal, Madhulika Sharma, Stephen C. Parnell, Lesya Zelenchuk, M. Cristine Charlesworth, Benjamin J. Madden, Kenneth L. Johnson, Daniel J. McCormick, Marie C. Hogan, Christopher J. Ward

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

The polycystin–1 (PC1), polycystin–2 (PC2) and fibrocystin proteins, the respective products of the PKD1, PKD2 and PKHD1 genes, are abundant in urinary exosome–like vesicles (ELVs) where they form the polycystin complex (PCC). ELVs are 100 nm diameter membrane vesicles shed into the urine by the cells lining the nephron. Using MS/MS analysis of ELVs from individuals with PKD1 mutations and controls, we show that in addition to the well-described GPS/GAIN cleavage event in PC1 at 3048 aa and the proprotein convertase cleavage (PPC) event in fibrocystin at 3616 aa, there are multiple other cleavage events in these proteins. The C–terminal 11 transmembrane portion of PC1 undergoes three cleavage events in vivo. The absence of peptides from the C–terminal cytoplasmic tail of fibrocystin implies a cleavage event close to its single TM domain prior to loading onto the ELVs. There is also evidence that the C–terminal tail of PC2 is also cleaved in ELVs. Native gel analysis of the PCC shows that the entire complex is > 2 MDa in size and that N–terminal GPS/GAIN cleaved PC1 and PPC cleaved fibrocystin ectodomains can be released under non-reducing conditions and resolve at 300 kDa. This paper shows that the three major human cystogene proteins are detectable in human urinary ELVs and that all three undergo post-translational proteolytic processing. Human urinary ELVs may be a useful source of material in the search for proteins that interact with the PCC.

Original languageEnglish (US)
Article number1500
JournalScientific reports
Volume10
Issue number1
DOIs
StatePublished - Dec 1 2020

ASJC Scopus subject areas

  • General

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