Identification of a New 5'-Noncoding Exon Region and Promoter Activity in Human N-Acetylglucosaminyltransferase III Gene

  • Kang, Bong-Seok ;
  • Kim, Yeon-Jeong ;
  • Shim, Jae-Kyoung ;
  • Song, Eun-Young ;
  • Park, Young-Guk ;
  • Lee, Young-Choon ;
  • Nam, Kyung-Soo ;
  • Kim, June-Ki ;
  • Lee, Tae-Kyun ;
  • Chung, Tae-Wha ;
  • Kim, Cheorl-Ho
  • Received : 1998.07.13
  • Accepted : 1998.09.03
  • Published : 1998.11.30

Abstract

In a previous paper (Kim et al., 1996a), the immediate 5' -flanking region and coding region of the human UDP-N -acetylglucosamine:-D-mannoside-1,4-Nacetylglucosaminyltransferase III (N-acetylglucosaminyitransferase- III; GnT-III) gene was reported, isolated and analyzed. Herein, we report on amplification of a new 5' -noncoding region of the GnT-III mRNA by single-strand ligation to single-stranded cDNA-PCR (5' -RACE PCR) using poly(A)+ RNA isolated from human fetal liver cells. A cDNA clone was obtained with 5' sequences (96 bp) that diverged seven nucleotides upstream from the ATG (+1) start codon. A concensus splice junction sequence, TCTCCCGCAG, was found immediately 5' to the position where the sequences of the cDNA diverged. The result suggested the presence of an intron in the 5' -noncoding region and that the cDNA was an incompletely reversetranscribed cDNA product derived from an mRNA containing a new noncoding exon. When mRNA expression of GnT-III in various human tissues and cancer cell lines was examined, Northern blot analysis indicated high expression levels of GnT-III in human fetal kidney and brain tissues, as well as for a number of leukemia and lymphoma cancer cell lines. Promoter activities of the 5' -flanking regions of exon 1 and the new noncoding region were measured in a human hepatoma cell line, HepG2, by luciferase assays. The 5'-flanking region of exon 1 was the most active, whilst that of exon 2 was inactive.

Keywords

D-N -acetylglucosamine:-D-mannoside-1,4-Nacetylglucosaminyltransferase III;Expression;Promoter