Characterization of human SHC p66 cDNA and its processed pseudogene mapping to Xq12-q13.1.
SHC is an adapter protein in the Ras-MAPkinase pathway that is involved in the regulation of cell growth and differentiation. The p46 and p52 isoforms are thought to be produced by the use of two alternative translation initiation sites in a 3.4-kb transcript from the SHCA gene, which maps to chromosome 1q21. The p66 isoform could be encoded by a different 3.8- or 2.8-kb transcript of the same gene or alternatively by a SHC-related gene. To characterize other putative genes coding for SHC-like proteins, primers from the 3' UTR of the SHCA gene were used to screen a yeast artificial chromosome (YAC) library by polymerase chain reaction (PCR). Two YAC clones, 20D11B and 36D1D, were isolated and used as probes for fluorescence in situ hybridization analysis. Both these probes hybridized to chromosome Xq12-q13.1. This novel SHC-related sequence was characterized by direct sequencing of vectorette library PCR products produced from clone 20D11B. A transcript of 3.2 kb that was 85% identical to the mouse Shc cDNA encoding the p66 isoform was identified. Sequence analysis demonstrated the presence of multiple stop codons identifying this isoform of SHC as a processed pseudogene. Using primers designed on the basis of the nucleotide sequence of the pseudogene, we have now amplified and sequenced a human cDNA that encodes the SHC p66 protein. Thus, we have characterized the human SHC p66 isoform cDNA and identified a processed SHC pseudogene that maps to chromosome Xq12-q13.1.
Harun
R B
RB
Molecular Medicine Unit, University of Leeds, St. James' University Hospital, United Kingdom.
Smith
K K
KK
Leek
J P
JP
Markham
A F
AF
Norris
A
A
Morrison
J F
JF
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002874:Chromosome Mapping; D018244:Chromosomes, Artificial, Yeast; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D008969:Molecular Sequence Data; D011506:Proteins / Q000235:genetics*; D011544:Pseudogenes; D012323:RNA Processing, Post-Transcriptional; D017386:Sequence Homology, Amino Acid; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014960:X Chromosome / Q000235:genetics*
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D017931:DNA Primers; D018076:DNA, Complementary; D011506:Proteins; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; C528034:Shc1 protein, mouse; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1