The cloning and preliminarily functional analysis of the human neurotrimin gene.
Proteins of the immunoglobulin superfamily (IgSF) are involved in a variety of specific cell-cell interactions in the developing nervous system. To identify and characterize new members of this protein family in human nervous system, we screen the human fetal brain cDNA library and isolate a full-length cDNA clone which contains a 1032 bp open reading frame encoding a protein of 344 amino acids. Sequence analysis reveals that it is a glycoprotein comprised of three C2-like immunoglobulin domains and is anchored to the plasma membrane via a post-translationally attached glycosyl-phosphatidylinositol (GPI) moiety. The protein shows high sequence similarity to the rat Ntm (97%), so we term it human neurotrimin (NTM). Northern blot analysis reveals that (HUMAN)NTM has three different transcripts with the length of 3.2 kb, 4.0 kb and 9.0 kb respectively. It has a wider expression pattern than that of (RAT) Ntm. Notably, the expression of NTM in fetal brain is higher than that in mature brain and is stronger in nervous tumors than that in normal brain tissues. We insert an HA epitope tag between the third Ig-like domain of NTM and the site of GPI attachment, then construct it into the eukaryotic expression vector pcDNA3.1+/Zeocin. The pcDNA3.1-HA-NTM is transfected into the Chinese Hamster Ovary (CHO) cells. The results demonstrate that HA-NTM is expressed on the surface of CHO cells and could strengthen the aggregation of CHO-NTM cells.
Liu
Jin
J
National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences, School of Basic Medicine, Peking Union Medical College, National Human Genome North Center, Beijing 100005, China.
Li
Guangtao
G
Peng
Xiaozhong
X
Liu
Bingyan
B
Yin
Bin
B
Tan
Xinyu
X
Fan
Ming
M
Fan
Wenhong
W
Qiang
Boqin
B
Yuan
Jiangang
J
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002449:Cell Aggregation / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods; D018388:Codon, Terminator / Q000235:genetics; D006224:Cricetinae; D017931:DNA Primers; D020224:Expressed Sequence Tags; D058851:GPI-Linked Proteins; D006801:Humans; D007136:Immunoglobulins / Q000235:genetics; D008969:Molecular Sequence Data; D019006:Neural Cell Adhesion Molecules / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D018388:Codon, Terminator; D017931:DNA Primers; D058851:GPI-Linked Proteins; D007136:Immunoglobulins; D019006:Neural Cell Adhesion Molecules; D012333:RNA, Messenger; C092530:neurotrimin