Ras mutation impairs epithelial barrier function to a wide range of nonelectrolytes.
Although ras mutations have been shown to affect epithelial architecture and polarity, their role in altering tight junctions remains unclear. Transfection of a valine-12 mutated ras construct into LLC-PK1 renal epithelia produces leakiness of tight junctions to certain types of solutes. Transepithelial permeability of D-mannitol increases sixfold but transepithelial electrical resistance increases >40%. This indicates decreased paracellular permeability to NaCl but increased permeability to nonelectrolytes. Permeability increases to D-mannitol (Mr 182), polyethylene glycol (Mr 4000), and 10,000-Mr methylated dextran but not to 2,000,000-Mr methylated dextran. This implies a "ceiling" on the size of solutes that can cross a ras-mutated epithelial barrier and therefore that the increased permeability is not due to loss of cells or junctions. Although the abundance of claudin-2 declined to undetectable levels in the ras-overexpressing cells compared with vector controls, levels of occludin and claudins 1, 4, and 7 increased. The abundance of claudins-3 and -5 remained unchanged. An increase in extracellular signal-regulated kinase-2 phosphorylation suggests that the downstream effects on the tight junction may be due to changes in the mitogen-activated protein kinase signaling pathway. These selective changes in permeability may influence tumorigenesis by the types of solutes now able to cross the epithelial barrier.
Mullin
James M
JM
The Lankenau Institute for Medical Research, Wynnewood, PA 19096, USA. mullinj@mlhs.org
Leatherman
James M
JM
Valenzano
Mary Carmen
MC
Huerta
Erika Rendon
ER
Verrechio
Jon
J
Smith
David M
DM
Snetselaar
Karen
K
Liu
Mantao
M
Francis
Mary Kay
MK
Sell
Christian
C
D000818:Animals; D001692:Biological Transport; D002455:Cell Division; D002460:Cell Line; D002463:Cell Membrane Permeability; D003911:Dextrans / Q000378:metabolism; D004594:Electrophysiology; D011905:Genes, ras; D007668:Kidney; D008565:Membrane Proteins / Q000378:metabolism; D009154:Mutation; D012965:Sodium Chloride / Q000378:metabolism; D014162:Transfection; D019459:Urothelium / Q000502:physiology*
D003911:Dextrans; D008565:Membrane Proteins; D012965:Sodium Chloride; C014392:carboxymethyl dextran