Characterization and tissue expression of a novel human gene npdc1☆
Introduction
Transcription factors in the basic-helix-loop-helix (bHLH) family are key regulators in development, particularly in cell-type determination, terminal differentiation, and sex determination (Yang et al., 2000). Within the last decade, several mammalian neural HLH proteins have been found to play critical roles in neural terminal differentiation, such as MASH-1 and -2 (Johnson et al., 1990, Horton et al., 1999), MATH-1 and -2 (Akazawa et al., 1995), NeuroD (Lee et al., 1995, Liu et al., 2000), neurogenin (Ma et al., 1996), HES-1 and -2 (Sasai et al., 1992, Ishibashi et al., 1993). These results promoted a lot of researchers to search for novel mammalian HLH factors which may be involved in the specific control of neural cell proliferation and differentiation.
Many new genes have been identified from human fetal liver during recent years with the progress of large-scale cDNA sequencing project in our lab (Yang et al., 1997, Wang et al., 1999, Zhang et al., 2000). Up to date, more than 18 000 expressed sequence tags (ESTs) have been analyzed to search the non-redundant GenBank database with the Blast program. Among these ESTs, an insert clone FLD2599 encodes a putative partial polypeptide of 305 amino acids, and a segment (LEEEIDSLAQELALKE) in amino-terminal of the polypeptide shows moderate homology with HLH domains in several known regulatory factors (e.g. amino acids 190–207 of Fos B and amino acids 276–285 of c-Jun). So this clone was selected and extended towards the 5′ end by PCR amplification of cDNA ends. This has led to obtain the entire coding sequence of a novel human gene, which shows high homology to the mouse npdc1 (mnpdc1) and so we term it hnpdc1 (human neural proliferation, differentiation and control, 1). The neural-specific mnpdc1 cDNA was isolated from postnatal mouse brain by using oligonucleotides derived from HLH recognition sequences as probes (Gallana et al., 1995). The protein mNPDC1 does not belong to any known HLH subfamily, although it contains an HLH-like domain. It down-regulated cell proliferation and its expression was maintained in growth-arrested and differentiated cells (Dupont et al., 1998). Here we report the sequence features of the cDNA and the predictive protein, gene identification, chromosomal assignment and tissue expression of hnpdc1.
Section snippets
Identification and cloning of hnpdc1
In our lab, a cDNA library of human fetal liver (second trimester) was selected for large-scale DNA sequencing to search novel genes with important functions. An on-line Blast search was performed against the public EST and non-redundant GenBank database with the EST sequence (Altschul et al., 1997). The clone FLD2599 susceptible of encoding a protein with HLH-like domain was picked up for further analysis.
5′-Rapid amplification of cDNA ends (RACE)
A new generation of SMART™ RACE cDNA Amplification Kit (CLONTECH) was used. MMLV reverse
Identification and cloning of hnpdc1
After sequencing the cDNA library of human fetal liver and searching the non-redundant GenBank database, a cDNA clone (No. FLD2599) susceptible of encoding protein with HLH-like domain and homologous to mnpdc1 cDNA was selected for further study. The insertion segment of this clone was 1283 bp (including 61 adenine bases at 3′ terminal) in size and included an open reading frame (ORF) of 918 nucleotides that lacked an initiation codon. Since the segment contains poly(A) addition signal and
Acknowledgements
This subject was partially supported by Chinese National Natural Sciences Foundation (30070177), Chinese National Natural Sciences Foundation Key Project (39730310), Chinese National Key Project of Basic Research and Chinese High-tech Program.
References (23)
- et al.
A mammalian helix-loop-helix factor structurally related to the product of Drosophila proneural gene atonal is a positive transcriptional regulator expressed in the developing nervous system
J. Biol. Chem.
(1995) - et al.
A cDNA encoding a pRb-bing protein with properties of the transcription factor E2F
Cell
(1992) - et al.
Correct coordination of neuronal differentiation events in ventral forebrain requires the bHLH factor MASH1
Mol. Cell. Neurosci.
(1999) Initiation of translation in prokaryotes and eukaryotes
Gene
(1999)- et al.
Identification of neurogenin, a vertebrate neuronal determination gene
Cell
(1996) - et al.
Identification and characterization of receptor for mammalian hepatopoietin that is homologous to yeast ERV1
J. Biol. Chem.
(1999) The retinoblastoma protein and the cell cycle control
Cell
(1995)- et al.
Characterization, chromosomal assignment, and tissue expression of a novel human gene belong to ARF GAP family
Genomics
(2000) - et al.
Gapped BLAST and PSI-BLAST: a new generation of protein database search programs
Nucleic Acids Res.
(1997) - et al.
Developmental pattern of expression of NPDC-1 and its interaction with E2F-1 suggest a role in the control of proliferation and differentiation of neural cells
J. Neurosci. Res.
(1998)
Identification of a neural-specific cDNA, NPDC-1, able to down-regulate cell proliferation and to suppress transformaion
Proc. Natl. Acad. Sci. USA
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Data deposition: The sequence reported in this paper has been deposited in the GenBank Accession (Number AF285836).