The cis-regulatory element Gsl5 is indispensable for proximal straight tubule cell-specific transcription of core 2 beta-1,6-N-acetylglucosaminyltransferase in the mouse kidney.
Gsl5 regulates the expression of a glycolipid and glycoproteins that contain the Le(X) epitope in the mouse kidney through tissue-specific transcriptional regulation of the core 2 beta-1,6-N-acetylglucosaminyltransferase (core 2 GnT) gene. The core 2 GnT gene has six exons and produces three alternatively spliced transcripts. Gsl5 regulates only the expression of the kidney-type mRNA, which is transcribed from the most 5'-upstream exon. By introducing a 159-kb bacterial artificial chromosome (BAC) clone that carries the mouse core 2 GnT gene and its 5'-upstream region into DBA/2 mice that carry a defective Gsl5 allele, we were able to rescue the deficient phenotype. The BAC clone was subsequently engineered to replace the core 2 GnT gene with the sequence of enhanced green fluorescent protein (EGFP) as a reporter by an inducible homologous recombination system in Escherichia coli. The transgenic mice derived from the modified BAC clone expressed EGFP in the kidney, which suggests that the candidate Gsl5 is in the 5'-upstream region of the core 2 GnT gene. Sequence analysis of the 5'-upstream regions of the BAC clone and DBA/2 genomic DNA revealed a candidate sequence for Gsl5 at about 5.5 kb upstream of exon 1. This sequence consisted of eight repeats of two GT-rich units in the wild-type mice, whereas it consisted of only one pair of GT-rich units with a minor modification in the DBA/2 mice. Transgenic mice produced with the EGFP reporter gene construct that included this candidate sequence expressed EGFP exclusively in the proximal straight tubular cells of the kidney. These results indicated that this unique repeat is indeed the Gsl5, and it is a cis-regulatory element responsible for proximal straight tubule cell-specific transcriptional regulation.
Sekine
Michiko
M
Department of Laboratory Animal Science, Tokyo Metropolitan Institute of Medical Science, Japan.
Taya
Chouji
C
Shitara
Hiroshi
H
Kikkawa
Yoshiaki
Y
Akamatsu
Noriko
N
Kotani
Masaharu
M
Miyazaki
Masao
M
Suzuki
Akemi
A
Yonekawa
Hiromichi
H
D000483:Alleles; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D022202:Chromosomes, Artificial, Bacterial / Q000378:metabolism; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004926:Escherichia coli / Q000378:metabolism; D005091:Exons; D017353:Gene Deletion; D015723:Gene Library; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000378:metabolism; D007668:Kidney / Q000378:metabolism* / Q000473:pathology; D007684:Kidney Tubules / Q000166:cytology*; D051379:Mice; D008811:Mice, Inbred DBA; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D008957:Models, Genetic; D008969:Molecular Sequence Data; D017351:N-Acetylglucosaminyltransferases / Q000378:metabolism / Q000502:physiology*; D010641:Phenotype; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D011995:Recombination, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014018:Tissue Distribution; D014157:Transcription Factors / Q000235:genetics* / Q000502:physiology*; D014158:Transcription, Genetic; D019076:Transgenes
D017931:DNA Primers; C508149:Gsl5 protein, mouse; D012333:RNA, Messenger; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D017351:N-Acetylglucosaminyltransferases; C053024:beta-1,3-galactosyl-O-glycosyl-glycoprotein beta-1,6-acetylglucosaminyl transferase