Cloning and sequencing of the 52K cathepsin D complementary deoxyribonucleic acid of MCF7 breast cancer cells and mapping on chromosome 11

Two lambda gt11 libraries containing complementary DNAs from human breast cancer MCF7 cells were screened by expression with monoclonal antibodies to the secreted 52K protein and with a 36-mer oligonucleotide derived from the N-terminal amino acid sequence of the secreted 52K protein. Four overlapping clones were sequenced, and found to be extensively homologous to the cathepsin D of normal human kidney, except for 5-point mutations resulting in one amino acid change (Ala to Val) in the profragment of cathepsin D. Northern blot analysis showed the 2.2 kilobase (kb) cathepsin D mRNA to be induced by estradiol in MCF7 cells and produced constitutively at high levels in the estrogen-receptor-negative BT20 cell line. A simple restriction pattern consistent with the restriction map of cathepsin D cDNA was obtained in Southern blot analysis of MCF7 cell DNA. In situ hybridization of the 52K-9 cDNA probe on normal lymphocytes assigned the 52K cathepsin D gene at the extremity of the short arm of chromosome 11, in the p15 band, close to the H-ras gene and in the region whose deletion increases the risk of invasive breast cancer. We conclude that the estrogen induced 52K protein has the same sequence as normal pro-cathepsin D and we propose that the 52K protein correspond to the only pro-cathepsin D expressed in MCF7 cells.     

Chromosomal localization of the mouse gene coding for the 68 kDa neurofilament subunit

The chromosomal localization of the mouse gene coding for the 68 kDa intermediate filament subunit of neurones (NF-L) was determined by in situ hybridization using specific 3H-labelled DNA probes. There is only one copy of the NF-L gene. The gene encoding NF-L is located on chromosome 14 region (D1-E1).     

Evidence for allelism of the recessive insertional mutation add and the dominant mouse mutation extra-toes (Xt)

A recessive mutant caused by insertional mutagenesis in transgenic mice has been detected in which the anterior part of the forelimb is disorganized. The morphology of the thumb is always altered and sometimes the adjacent finger has an extra phalanx. This phenotype suggests that a body plan gene is affected. We have named the mutation add (anterior digit-pattern deformity). Using the cloned DNA from the flanking region of the integrated transgene, add has been mapped close to the centromere of chromosome 13. This position links add to a genetically mapped locus called extra-toes (Xt). The phenotype of the double-mutant add/Xt as well as the molecular analysis suggest that add and Xt are allelic.     

Molecular cloning of rat and human type IX collagen cDNA and localization of the alpha 1(IX) gene on the human chromosome 6

Type IX collagen is found in hyaline cartilage, where it is associated with type II collagen in quarter-staggered collagen fibrils. Chicken type IX collagen has been extensively characterized and shown to contain molecules with three triple-helical domains, interspersed with non-triple-helical sequences. The molecule contains three, genetically distinct, subunits and one of these subunits carries a covalently bound glycosaminoglycan side chain. In the present report, we describe for the first time the primary structure of mammalian type IX collagen chains, based on cloning and sequencing of cDNA from rat and human cDNA libraries. The results suggest that mammalian alpha 1(IX) chains have the same multi-domain structure as the avian protein. We also demonstrate, by in situ hybridization of chromosome spreads, that the human alpha 1(IX) collagen gene is located on the long arm of chromosome 6. The cloning of human type IX collagen cDNA provides a probe for molecular studies of human chondrodysplasias that may involve abnormalities in this extracellular collagen-proteoglycan.     

Chromosomal localization of the mouse gene coding for vimentin

The chromosomal location of the mouse gene coding for vimentin, one of the intermediate filament subunits, was determined by in situ hybridization using specific H3-labelled DNA probes. There is only one copy of the vimentin gene and it is located on chromosome 2 region A2.     

Reactivity of the human monoclonal antibody 33G2 with repeated sequences of three distinct Plasmodium falciparum antigens

The human mAb 33G2 has high capacity to inhibit in vitro invasion of erythrocytes by Plasmodium falciparum merozoites and, thus, is of special interest with regard to protective immunity against the parasite. In order to obtain more information about asexual blood stage Ag of P. falciparum that are seen by this antibody, material from synchronized P. falciparum cultures was studied by immunofluorescence, immunoelectron microscopy, and immunoblotting. Reactivity was mainly confined to the membrane of infected erythrocytes. Soon after merozoite invasion the antibody stained the erythrocyte membrane. This membrane-associated staining faded during intracellular development of the parasites. Beginning about 18 h after invasion, a dotted pattern appeared which increased in strength with time and persisted to schizont rupture. Pf155/RESA was the major Ag recognized in immunoblots of parasites collected throughout the entire erythrocytic cycle, although other polypeptides also bound the antibody. Among these was a 260-kDa polypeptide found in late trophozoites and schizonts. The specificity of the antibody was analyzed with synthetic peptides corresponding to repeated sequences in the P. falciparum Ag Pf155/RESA, Pf11.1, and Ag332. Synthetic peptides related to Ag332 were the most efficient inhibitors of antibody binding in immunofluorescence studies and cell ELISA. A beta-galactosidase-Ag332 fusion protein was also efficient in reversing reinvasion inhibition caused by 33G2. These results define a family of cross-reactive P. falciparum Ag recognized by mAb 33G2 and suggest that Ag332 was its original target.     

Ultrastructure of the spermatozoid of Centrorhynchus milvus Ward 1956 (Paleacanthocephala, Polymorphidae)]

The Centrorhynchus milvus spermatozoon is a filiform cell. The free part of the flagellum measure 2 mu and is situated in front of the gamete. The centriole is no more visible but an axial tubule formation is found at the anterior extremity of the flagellum. This Polymorphidae spermatozoon shows a reversed anatomy like the one we described for the first time with an acanthocephala wich belongs to the Rhadinorhynchidae family.     

Cone-shaped HIV-1 capsids are transported through intact nuclear pores

1. Download : Download high-res image (250KB)
2. Download : Download full-size image