Quantitative mass spectrometry reveals a role for the GTPase Rho1p in actin organization on the peroxisome membrane.
We have combined classical subcellular fractionation with large-scale quantitative mass spectrometry to identify proteins that enrich specifically with peroxisomes of Saccharomyces cerevisiae. In two complementary experiments, isotope-coded affinity tags and tandem mass spectrometry were used to quantify the relative enrichment of proteins during the purification of peroxisomes. Mathematical modeling of the data from 306 quantified proteins led to a prioritized list of 70 candidates whose enrichment scores indicated a high likelihood of them being peroxisomal. Among these proteins, eight novel peroxisome-associated proteins were identified. The top novel peroxisomal candidate was the small GTPase Rho1p. Although Rho1p has been shown to be tethered to membranes of the secretory pathway, we show that it is specifically recruited to peroxisomes upon their induction in a process dependent on its interaction with the peroxisome membrane protein Pex25p. Rho1p regulates the assembly state of actin on the peroxisome membrane, thereby controlling peroxisome membrane dynamics and biogenesis.
Marelli
Marcello
M
Institute for Systems Biology, Seattle, WA 98103, USA.
Smith
Jennifer J
JJ
Jung
Sunhee
S
Yi
Eugene
E
Nesvizhskii
Alexey I
AI
Christmas
Rowan H
RH
Saleem
Ramsey A
RA
Tam
Yuen Yi C
YY
Fagarasanu
Andrei
A
Goodlett
David R
DR
Aebersold
Ruedi
R
Rachubinski
Richard A
RA
Aitchison
John D
JD
D000199:Actins / Q000032:analysis / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000378:metabolism; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D008962:Models, Theoretical; D000074401:Peroxins; D020675:Peroxisomes / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
D000199:Actins; D008565:Membrane Proteins; C475282:PEX25 protein, S cerevisiae; D000074401:Peroxins; D029701:Saccharomyces cerevisiae Proteins; D020558:GTP Phosphohydrolases; C051302:RHO1 protein, S cerevisiae; D020741:rho GTP-Binding Proteins