PMID- 31694916
OWN - NLM
STAT- MEDLINE
DCOM- 20200713
LR  - 20240422
IS  - 1083-351X (Electronic)
IS  - 0021-9258 (Print)
IS  - 0021-9258 (Linking)
VI  - 294
IP  - 50
DP  - 2019 Dec 13
TI  - Primary cilia and the exocyst are linked to urinary extracellular vesicle 
      production and content.
PG  - 19099-19110
LID - 10.1074/jbc.RA119.009297 [doi]
AB  - The recently proposed idea of "urocrine signaling" hypothesizes that small 
      secreted extracellular vesicles (EVs) contain proteins that transmit signals to 
      distant cells. However, the role of renal primary cilia in EV production and 
      content is unclear. We previously showed that the exocyst, a highly conserved 
      trafficking complex, is necessary for ciliogenesis; that it is present in human 
      urinary EVs; that knockdown (KD) of exocyst complex component 5 (EXOC5), a 
      central exocyst component, results in very short or absent cilia; and that human 
      EXOC5 overexpression results in longer cilia. Here, we show that compared with 
      control Madin-Darby canine kidney (MDCK) cells, EXOC5 overexpression increases 
      and KD decreases EV numbers. Proteomic analyses of isolated EVs from EXOC5 
      control, KD, and EXOC5-overexpressing MDCK cells revealed significant alterations 
      in protein composition. Using immunoblotting to specifically examine the 
      expression levels of ADP-ribosylation factor 6 (ARF6) and EPS8-like 2 (EPS8L2) in 
      EVs, we found that EXOC5 KD increases ARF6 levels and decreases EPS8L2 levels, 
      and that EXOC5 overexpression increases EPS8L2. Knockout of intraflagellar 
      transport 88 (IFT88) confirmed that the changes in EV number/content were due to 
      cilia loss: similar to EXOC5, the IFT88 loss resulted in very short or absent 
      cilia, decreased EV numbers, increased EV ARF6 levels, and decreased Eps8L2 
      levels compared with IFT88-rescued EVs. Compared with control animals, urine from 
      proximal tubule-specific EXOC5-KO mice contained fewer EVs and had increased ARF6 
      levels. These results indicate that perturbations in exocyst and primary cilia 
      affect EV number and protein content.
FAU - Zuo, Xiaofeng
AU  - Zuo X
AD  - Department of Medicine, Medical University of South Carolina, Charleston, South 
      Carolina 29425.
FAU - Kwon, Sang-Ho
AU  - Kwon SH
AD  - Department of Cellular Biology and Anatomy, Augusta University, Augusta, Georgia 
      30912.
FAU - Janech, Michael G
AU  - Janech MG
AD  - Department of Biology, College of Charleston, Charleston, South Carolina 29424.
FAU - Dang, Yujing
AU  - Dang Y
AD  - Department of Medicine, Medical University of South Carolina, Charleston, South 
      Carolina 29425.
FAU - Lauzon, Steven D
AU  - Lauzon SD
AD  - Department of Public Health Sciences, Medical University of South Carolina, 
      Charleston, South Carolina 29425.
FAU - Fogelgren, Ben
AU  - Fogelgren B
AD  - Department of Anatomy, Biochemistry, and Physiology, University of Hawaii at 
      Manoa, Honolulu, Hawaii 96813.
FAU - Polgar, Noemi
AU  - Polgar N
AUID- ORCID: 0000-0002-8400-162X
AD  - Department of Anatomy, Biochemistry, and Physiology, University of Hawaii at 
      Manoa, Honolulu, Hawaii 96813.
FAU - Lipschutz, Joshua H
AU  - Lipschutz JH
AD  - Department of Medicine, Medical University of South Carolina, Charleston, South 
      Carolina 29425 Lipschut@musc.edu.
AD  - Department of Medicine, Ralph H. Johnson Veterans Affairs Medical Center, 
      Charleston, South Carolina 29425.
LA  - eng
GR  - R01 DK117308/DK/NIDDK NIH HHS/United States
GR  - P30 DK074038/DK/NIDDK NIH HHS/United States
GR  - P30 GM131944/GM/NIGMS NIH HHS/United States
GR  - R01 DK120510/DK/NIDDK NIH HHS/United States
GR  - K01 DK087852/DK/NIDDK NIH HHS/United States
GR  - R03 DK100738/DK/NIDDK NIH HHS/United States
GR  - P20 GM113134/GM/NIGMS NIH HHS/United States
GR  - P20 GM103457/GM/NIGMS NIH HHS/United States
GR  - UL1 TR001450/TR/NCATS NIH HHS/United States
GR  - I01 BX000820/BX/BLRD VA/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
DEP - 20191106
PL  - United States
TA  - J Biol Chem
JT  - The Journal of biological chemistry
JID - 2985121R
RN  - 0 (ADP-Ribosylation Factor 6)
RN  - 0 (EXOC5 protein, human)
RN  - 0 (Vesicular Transport Proteins)
RN  - EC 3.6.5.2 (ARF6 protein, human)
RN  - EC 3.6.5.2 (Arf6 protein, mouse)
SB  - IM
MH  - ADP-Ribosylation Factor 6
MH  - Animals
MH  - Cells, Cultured
MH  - Cilia/*metabolism
MH  - Dogs
MH  - *Exocytosis
MH  - Extracellular Vesicles/*metabolism
MH  - Humans
MH  - Kidney/*metabolism
MH  - Madin Darby Canine Kidney Cells/metabolism
MH  - Mice
MH  - Mice, Knockout
MH  - Vesicular Transport Proteins/deficiency/*metabolism
PMC - PMC6916495
OTO - NOTNLM
OT  - cilia
OT  - exocytosis
OT  - extracellular vesicles
OT  - kidney
OT  - protein export
COIS- The authors declare that they have no conflicts of interest with the contents of 
      this article
EDAT- 2019/11/07 06:00
MHDA- 2020/07/14 06:00
PMCR- 2020/12/13
CRDT- 2019/11/08 06:00
PHST- 2019/05/09 00:00 [received]
PHST- 2019/10/29 00:00 [revised]
PHST- 2019/11/07 06:00 [pubmed]
PHST- 2020/07/14 06:00 [medline]
PHST- 2019/11/08 06:00 [entrez]
PHST- 2020/12/13 00:00 [pmc-release]
AID - S0021-9258(20)30824-3 [pii]
AID - RA119.009297 [pii]
AID - 10.1074/jbc.RA119.009297 [doi]
PST - ppublish
SO  - J Biol Chem. 2019 Dec 13;294(50):19099-19110. doi: 10.1074/jbc.RA119.009297. Epub 
      2019 Nov 6.