Urea movement across mouse colonic plasma membranes is mediated by UT-A urea transporters.
OBJECTIVE
Urea is a major nitrogen source for commensal bacteria that inhabit the large intestine. UT-A urea transporters mediate urea movement across plasma membranes. The aim of this study was to determine whether UT-A proteins are expressed in the mouse colon and, if so, whether they have a functional role in transcellular urea transport.


METHODS
Mouse colonic UT-A transporters were investigated with Northern blot analysis, immunoblotting, immunolocalization, and refractive light flux experiments.


RESULTS
Northern blot analysis showed that 4 UT-A transcripts were present in mouse colon. Two peptide-targeted polyclonal antibodies showed the presence of UT-A immunoreactive proteins in mouse colon. Antiserum ML446 targeted to the N-terminus of mouse UT-A1 detected proteins of 34 and 48 kilodaltons. Antiserum ML194 targeted to the C-terminus of mouse UT-A1 detected proteins of 48, 75, and 100 kilodaltons. Immunolocalization studies using ML446 showed the presence of UT-A proteins in cells throughout the colonic crypts. ML194 specifically stained cells located in the proliferative and stem regions of the lower portion of colonic crypts. Differential centrifugation and immunoblotting of colonic epithelia showed that UT-A proteins were present in plasma membrane-enriched fractions. Refractive light flux experiments using colonic plasma membrane vesicles showed a significant urea flux, which was completely inhibited by the UT-A inhibitor phloretin.


CONCLUSIONS
Functional UT-A transporters are expressed in the plasma membranes of mouse colon, indicating that these proteins may play a key role in host/bacterial interaction.
Stewart
Gavin S
GS
School of Biological Sciences, University of Manchester, England.
Fenton
Robert A
RA
Thévenod
Frank
F
Smith
Craig P
CP
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D015152:Blotting, Northern; D002462:Cell Membrane / Q000378:metabolism; D002498:Centrifugation; D003106:Colon / Q000378:metabolism*; D007106:Immune Sera; D015151:Immunoblotting; D007158:Immunologic Techniques; D007413:Intestinal Mucosa / Q000378:metabolism; D007668:Kidney / Q000378:metabolism; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008871:Microvilli / Q000378:metabolism; D010693:Phloretin / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D014508:Urea / Q000378:metabolism*; D000096982:Urea Transporters
D007106:Immune Sera; D049990:Membrane Transport Modulators; D026901:Membrane Transport Proteins; D010693:Phloretin; D012333:RNA, Messenger; D014508:Urea; D000096982:Urea Transporters