Metallothionein isoform 3 and proximal tubule vectorial active transport.
BACKGROUND
Metallothionein isoform 3 (MT-3) is expressed in the proximal tubule cells of the human kidney. The goal of the present study was to further characterize the basal expression of MT-3 in the proximal tubule and to determine if MT-3 participates in the maintenance of proximal tubule cell function.


METHODS
Expression of MT-3 mRNA was determined in the intact proximal tubule using microdissection and reverse transcription-polymerase chain reaction (RT-PCR). Basal expression of MT-3 mRNA and protein was determined in cultured human proximal tubule (HPT) cells and an immortalized proximal tubular cell line, HK-2 cells, using RT-PCR and immunoblotting. The MT-3 gene was stably transfected into the HK-2 cell line using the pcDNA3.1/Hygro (+) vector.


RESULTS
MT-3 mRNA was detected in the proximal tubule of the in situ kidney with relative expression in excess to that of the beta-actin housekeeping gene. The mortal HPT cells were shown to express both MT-3 mRNA and protein and to form domes, while immortal HK-2 cells were shown to have no expression of MT-3 mRNA and protein nor to form domes. The stable transfection of MT-3 in HK-2 restored MT-3 expression and dome formation to the HK-2 cells.


CONCLUSIONS
MT-3 mRNA is present in the human proximal tubule, and MT-3 expression is involved in the transport function of a human renal cell line that retains properties of the proximal tubule.
Kim
Doyeob
D
Department of Surgery, Program in Genetics and Developmental Biology, Robert C. Byrd Health Sciences Center, West Virginia University, Morgantown, West Virginia 26506-9251, USA.
Garrett
Scott H
SH
Sens
Mary Ann
MA
Somji
Seema
S
Sens
Donald A
DA
D000368:Aged; D001693:Biological Transport, Active / Q000502:physiology*; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D005260:Female; D015870:Gene Expression / Q000502:physiology; D006801:Humans; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000378:metabolism*; D008138:Loop of Henle / Q000166:cytology / Q000378:metabolism; D008297:Male; D008668:Metallothionein / Q000235:genetics / Q000378:metabolism*; D000090023:Metallothionein 3; D008875:Middle Aged; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D014162:Transfection
D000090023:Metallothionein 3; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; D008668:Metallothionein