The complete 1,210-amino acid sequence of the human epidermal growth factor (EGF) receptor precursor, deduced from cDNA clones derived from placental and A431 carcinoma cells, reveals close similarity between the entire predicted v-erb-B mRNA oncogene product and the receptor transmembrane and cytoplasmic domains. A single transmembrane region of 23 amino acids separates the extracellular EGF binding and cytoplasmic domains. The receptor gene is amplified and apparently rearranged in A431 cells, generating a truncated 2.8-kilobase mRNA which encodes only the extracellular EGF binding domain.

We have deduced the entire 1,370-amino-acid sequence of the human insulin receptor precursor from a single complementary DNA clone. The precursor starts with a 27-amino-acid signal sequence, followed by the receptor alpha-subunit, a precursor processing enzyme cleavage site, then the beta-subunit containing a single 23-amino-acid transmembrane sequence. There are sequence homologies to human epidermal growth factor receptor and the members of the src family of oncogene products.

No abstract

Structural features of v‐kit, the oncogene of HZ4 feline sarcoma virus, suggested that this gene arose by transduction and truncation of cellular sequences. Complementary DNA cloning of the human proto‐oncogene coding for a receptor tyrosine kinase confirmed this possibility: c‐kit encodes a transmembrane glycoprotein that is structurally related to the receptor for macrophage growth factor (CSF‐1) and the receptor for platelet‐derived growth factor. The c‐kit gene is widely expressed as a single, 5‐kb transcript, and it is localized to human chromosome 4 and to mouse chromosome 5. A c‐kit peptide antibody permitted the identification of a 145,000 dalton c‐kit gene product that is inserted in the cellular plasma membrane and is capable of self‐phosphorylation on tyrosine residues in both human glioblastoma cells and transfected mouse fibroblasts. Our results suggest that p145c‐kit functions as a cell surface receptor for an as yet unidentified ligand. Furthermore, carboxy‐ and amino‐terminal truncations that occurred during the viral transduction process are likely to have generated the transformation potential of v‐kit.

In vivo transduction of nondividing cells by human immunodeficiency virus type 1 (HIV-1)-based vectors results in transgene expression that is stable over several months. However, the use of HIV-1 vectors raises concerns about their safety. Here we describe a self-inactivating HIV-1 vector with a 400-nucleotide deletion in the 3′ long terminal repeat (LTR). The deletion, which includes the TATA box, abolished the LTR promoter activity but did not affect vector titers or transgene expression in vitro. The self-inactivating vector transduced neurons in vivo as efficiently as a vector with full-length LTRs. The inactivation design achieved in this work improves significantly the biosafety of HIV-derived vectors, as it reduces the likelihood that replication-competent retroviruses will originate in the vector producer and target cells, and hampers recombination with wild-type HIV in an infected host. Moreover, it improves the potential performance of the vector by removing LTR sequences previously associated with transcriptional interference and suppression in vivo and by allowing the construction of more-stringent tissue-specific or regulatable vectors.

No abstract

Nerve growth factor (NGF) is thought to have a profound effect on the development and maintenance of sympathetic and embryonic sensory neurones (see refs 1-3 for review). NGF activity isolated from the male mouse submaxillary gland (MSG) consists of three types of subunits, alpha, beta and gamma, which specifically interact to form a 7S, approximately 130,000-molecular weight (Mr) complex. The 7S complex contains two identical 118-amino acid beta-chains, which are solely responsible for the nerve growth-stimulating activity of NGF. While NGF is found in almost all vertebrates, most research has focused on murine NGF, as the mouse male submaxillary gland contains higher levels of this polypeptide than other tissues. Even so, beta-NGF comprises only approximately 0.1% of the protein in this small gland, which has made the study of this polypeptide difficult. The amino acid sequence of the mouse NGF beta-chain has been determined and some information has been obtained regarding the size of a mouse precursor molecule, pro-beta-NGF, but little was known about the structure and relatedness of beta-NGF from other vertebrates. Here we describe the isolation of mouse beta-NGF complementary DNA (cDNA) and present its nucleotide sequence, which predicts a prepro-beta-NGF molecule of Mr 27,000 (27K) and a pro-beta-NGF molecule of Mr 25K. We have used the mouse beta-NGF cDNA clone to isolate the human beta-NGF gene, the coding regions of which are highly homologous to the mouse prepro-beta-NGF nucleotide and amino acid sequences.

Cloning of the insulin receptor cDNA has earlier revealed the existence of two alternative forms of the receptor differing by the presence or absence of 12 amino acids near the C‐terminus of the receptor alpha‐subunit. This insert has been shown by others to be encoded by a discrete exon, and alternative splicing of this exon leads to tissue‐specific expression of two receptor isoforms. We have studied the functional significance of the receptor isoforms and have confirmed that they are generated by alternative splicing. When cDNAs encoding the two forms of the insulin receptors are expressed in Rat 1 cells, the receptor lacking the insert (HIR‐A) has a significantly higher affinity for insulin than the receptor with the insert (HIR‐B). This difference in affinity is maintained when insulin binding activity is assayed in solution using detergent solubilized, partially purified receptors. These data, combined with the tissue specificity of HIR‐A and HIR‐B expression, suggest that alternative splicing may result in the modulation of insulin metabolism or responsiveness by different tissues.