L. Tremblay et R. Beliveau, TYROSINE PROTEIN-PHOSPHORYLATION IN PLASMA-MEMBRANES OF RAT-KIDNEY CORTEX, The American journal of physiology, 267(3), 1994, pp. 60000415-60000422
The endogenous tyrosine protein kinase activity (TPKA) associated with
brush-border (BBM) and basolateral (BLM) membranes of rat kidney cort
ex was studied with an anti-phosphotyrosine monoclonal antibody (PY20)
. Distinct major phosphotyrosine-containing proteins were associated w
ith BBM (50, 54, and 120 kDa) and BLM (37, 90, 130, and 170 kDa). For
both plasma membranes, tyrosine phosphorylation leveled off after 10 m
in of incubation. Endogenous phosphotyrosine-specific protein phosphat
ases (PTPases) were active in both membranes, since the presence of so
dium vanadate or ammonium molybdate, which are inhibitors of PTPases,
was essential to detect endogenous phosphorylation. Substrates and/or
tyrosine protein kinases (TPKs) seem to be differently distributed in
these plasma membranes, since phosphorylation of endogenous substrates
in BLM and BBM was differently sensitive to competitive inhibitors of
TPKs. Moreover, insulin- and insulin-like growth factor I-stimulated
tyrosine phosphorylation of a 90-kDa substrate was only observed in so
lubilized BLM proteins. However, similar p60(v-src)-related TPKs appea
r to be present in the BBM and BLM, since an antibody raised against p
60(v-src) recognized proteins of 52, 58, and 75 kDa by immunoblotting
and could immunoprecipitate the TPKs associated with both plasma membr
anes. These data provide evidence that the endogenous tyrosine protein
phosphorylation observed in the ELM is catalyzed by nonreceptor TPKs
as well as receptor TPKs, whereas that observed in the BBM is exclusiv
ely due to nonreceptor TPKs.