Faculty Opinions recommendation of Human gene organization driven by the coordination of replication and transcription.

2007 ◽  
Author(s):  
Joel Huberman
Keyword(s):  
Human Gene ◽  
Gene Organization

scholarly journals Molecular cloning, gene organization and expression of the human UDP-GalNAc:Neu5Acalpha2-3Galbeta-R beta1,4-N-acetylgalactosaminyltransferase responsible for the biosynthesis of the blood group Sda/Cad antigen: evidence for an unusual extended cytoplasmic domain

2003 ◽  
Vol 373 (2) ◽  
pp. 369-379 ◽  
Author(s):  
Maria-Dolores MONTIEL ◽  
Marie-Ange KRZEWINSKI-RECCHI ◽  
Philippe DELANNOY ◽  
Anne HARDUIN-LEPERS
Keyword(s):  
Amino Acid ◽  
Catalytic Domain ◽  
Human Gene ◽  
Short Form ◽  
Cytoplasmic Tail ◽  
Gene Organization ◽  
Sialic Acid Residue ◽  
Chromosome 17 ◽  
Soluble Form ◽  
Amino Acid Residues

The human Sda antigen is formed through the addition of an N-acetylgalactosamine residue via a β1,4-linkage to a sub-terminal galactose residue substituted with an α2,3-linked sialic acid residue. We have taken advantage of the previously cloned mouse cDNA sequence of the UDP-GalNAc:Neu5Acα2-3Galβ-R β1,4-N-acetylgalactosaminyltransferase (Sda β1,4GalNAc transferase) to screen the human EST and genomic databases and to identify the corresponding human gene. The sequence spans over 35 kb of genomic DNA on chromosome 17 and comprises at least 12 exons. As judged by reverse transcription PCR, the human gene is expressed widely since it is detected in various amounts in almost all cell types studied. Northern blot analysis indicated that five Sda β1,4GalNAc transferase transcripts of 8.8, 6.1, 4.7, 3.8 and 1.65 kb were highly expressed in colon and to a lesser extent in kidney, stomach, ileum and rectum. The complete coding nucleotide sequence was amplified from Caco-2 cells. Interestingly, the alternative use of two first exons, named E1S and E1L, leads to the production of two transcripts. These nucleotide sequences give rise potentially to two proteins of 506 and 566 amino acid residues, identical in their sequence with the exception of their cytoplasmic tail. The short form is highly similar (74% identity) to the mouse enzyme whereas the long form shows an unusual long cytoplasmic tail of 66 amino acid residues that is as yet not described for any other mammalian glycosyltransferase. Upon transient transfection in Cos-7 cells of the common catalytic domain, a soluble form of the protein was obtained, which catalysed the transfer of GalNAc residues to α2,3-sialylated acceptor substrates, to form the GalNAcβ1-4[Neu5Acα2-3]Galβ1-R trisaccharide common to both Sda and Cad antigens.

Molecular cloning, gene organization and expression of rainbow trout (Oncorhynchus mykiss) inducible nitric oxide synthase (iNOS) gene

2001 ◽  
Vol 358 (3) ◽  
pp. 747-755 ◽  
Author(s):  
Tiehui WANG ◽  
Mike WARD ◽  
Peter GRABOWSKI ◽  
Christopher J. SECOMBES
Keyword(s):  
Nitric Oxide ◽  
Inducible Nitric Oxide Synthase ◽  
Reverse Transcription ◽  
Human Gene ◽  
Gene Organization ◽  
Inos Expression ◽  
Rt Pcr ◽  
Inos Gene ◽  
Oxide Synthase ◽  
Inducible Nitric Oxide

A full-length inducible nitric oxide synthase (iNOS) gene has been sequenced for the first time outside the mammals, and the gene organization compared with that already determined for human iNOS. While there are some differences from the human gene, overall the exons show remarkable conservation in sequence and organization. As in human, the trout iNOS gene has 27 exons, with 18 of the trout exons being identical in size with the equivalent human exons. The cofactor-binding domains are found in the same exons and in some cases are absolutely conserved. Differences include the start of the ORF in exon 3 instead of exon 2, resulting in a deletion at the 5′ end of the trout iNOS protein. Exon 27 also shows a large difference in size and although the trout exon is larger this is due to the length of the 3′-UTR. Several non-mammalian features are notable, and include a conserved potential glycosylation site in chicken and fish, and an insertion at the boundary of exons 20 and 21 in fish. The intron sizes in trout were generally much smaller than in human iNOS, making the trout iNOS gene approximately half the size of the human gene. Analysis of RNA secondary structure revealed two regions with complementarity, which could interfere with reverse transcription. Using a trout fibroblast cell line (RTG-2 cells), it was shown by reverse transcriptase (RT)-PCR that virus infection was a good inducer of iNOS expression. However, when using a combination of Superscript™ II for reverse transcription and primers at the 5′ end of the gene only very weak products were amplified, in contrast with the situation when primers at the 3′ end of the gene were used, or ThermoScript™-derived cDNA was used. The impact of such results on RT-PCR analysis of iNOS expression in trout is discussed.

scholarly journals Human gene organization driven by the coordination of replication and transcription

2007 ◽  
Vol 17 (9) ◽  
pp. 1278-1285 ◽  
Author(s):  
M. Huvet ◽  
S. Nicolay ◽  
M. Touchon ◽  
B. Audit ◽  
Y. d'Aubenton-Carafa ◽  
...  
Keyword(s):  
Human Gene ◽  
Gene Organization

The Influence of Glycosylation on the Catalytic and Fibrinolytic Properties of Pro-Urokinase

1992 ◽  
Vol 68 (05) ◽  
pp. 539-544 ◽  
Author(s):  
Catherine Lenich ◽  
Ralph Pannell ◽  
Jack Henkin ◽  
Victor Gurewich
Keyword(s):  
Escherichia Coli ◽  
Mammalian Cell ◽  
Human Gene ◽  
Culture Media ◽  
Mammalian Cell Culture ◽  
Glycosylation Site ◽  
Clot Lysis ◽  
Cell Culture Media ◽  
E Coli ◽  
The Uk

SummaryWe previously found that human pro-UK expressed in Escherichia coli is more active in fibrinolysis than recombinant human pro-UK obtained from mammalian cell culture media. To determine whether this difference is related to the lack of glycosylation of the E. coli product, we compared the activity of E. coli-derived pro-UK [(-)pro-UK] with that of a glycosylated pro-UK [(+)pro-UK] and of a mutant of pro-UK missing the glycosylation site at Asn-302 [(-) (302) pro-UK]. The latter two pro-UKs were obtained by expression of the human gene in a mammalian cell. The nonglycosylated pro-UKs were activated by plasmin more efficiently (≈2-fold) and were more active in clot lysis (1.5-fold) than the (+)pro-UK. Similarly, the nonglycosylated two-chain derivatives (UKs) were more active against plasminogen and were more rapidly inactivated by plasma inhibitors than the (+)UK.These findings indicate that glycosylation at Asn-302 influences the activity of pro-UK/UK and could be the major factor responsible for the enhanced activity of E. coli-derived pro-UK.

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