PubMed Health⌕ Search

Biomedical subjects

J A Engelman

Publications and source records attributed to J A Engelman.

26 records · Page 2Linked to original sources
PubMed1998

Chromosomal localization, genomic organization, and developmental expression of the murine caveolin gene family (Cav-1, -2, and -3). Cav-1 and Cav-2 genes map to a known tumor suppressor locus (6-A2/7q31).

Caveolins (Cav-1, -2, and -3) are a gene family of cytoplasmic membrane-anchored scaffolding proteins that: (i) help to sculpt caveolae membranes from the plasma membrane proper; and (ii) participate in the sequestration of inactive signaling molecules. In the adult, caveolin-1 and -2 are co-expressed and are most abundant in type I pneumocytes, endothelia, fibroblastic cells and adipocytes, while the expression of caveolin-3 is restricted to striated muscle cells. However, little is known regarding the genomic organization and developmental expression of the caveolin gene family. Here, using the mouse as a model system, we examine the chromosomal localization, the detailed intron-exon organization, and developmental expression pattern of the caveolin gene family. cDNAs encoding caveolin-1, -2, and -3 were used as probes to isolate murine genomic clones containing these genes. Fluorescence in situ hybridization (FISH) analysis using these genomic clones as probes reveals that all three caveolin genes are localized to murine chromosome 6. Specifically, caveolin-1 and -2 co-localize to chromosomal region 6-A2, while caveolin-3 is located within the chromosomal region 6-E1. Searches of the NCBI Human/Mouse Homology map indicate that murine region 6-A2 corresponds to human chromosome 7q31. As this region (6-A2/7q31) is the site of an as yet unidentified tumor suppressor gene(s), our mapping studies clearly define caveolin-1 and caveolin-2 as candidate genes that may be deleted at these loci. All three caveolin genes show similar intron-exon organization, with the last exon of each gene encoding the bulk of the known caveolin functional domains. The boundary position of the last exon is essentially identical in all three caveolin genes, suggesting that they may have arisen through gene duplication events. Developmentally, all three caveolins were expressed late during mouse embryogenesis as assessed by Northern and Western blot analysis. We examined the localization of the caveolin proteins in sections of day 16 mouse embryos using a well-characterized panel of antibody probes. Caveolin-1 and -2 were most abundantly expressed in the developing lung parenchyma, while caveolin-3 was most abundantly expressed in developing tissues that consist primarily of skeletal muscle cells. As the expression of all three caveolins in the adult is highest in terminally differentiated cell types, this is consistent with the idea that caveolins may be viewed as late markers of differentiation during embryogenesis.

Amino Acid Sequence↗
PubMed1998

Caveolin-mediated regulation of signaling along the p42/44 MAP kinase cascade in vivo. A role for the caveolin-scaffolding domain.

The p42/44 mitogen-activated protein (MAP)-kinase cascade is a well-established signal transduction pathway that is initiated at the cell surface and terminates within the nucleus. More specifically, receptor tyrosine kinases can indirectly activate Raf, which in turn leads to activation of MEK and ERK and ultimately phosphorylation of Elk, a nuclear transcription factor. Recent reports have suggested that some members of p42/44 MAP kinase cascade can be sequestered within plasmalemmal caveolae in vivo. For example, morphological studies have directly shown that ERK-1/2 is concentrated in plasma membrane caveolae in vivo using immunoelectron microscopy. In addition, constitutive activation of the p42/44 MAP kinase cascade is sufficient to reversibly down-regulate caveolin-1 mRNA and protein expression. However, the functional relationship between the p42/44 MAP kinase cascade and caveolins remains unknown. Here, we examine the in vivo role of caveolins in regulating signaling along the MAP kinase cascade. We find that co-expression with caveolin 1 dramatically inhibits signaling from EGF-R, Raf, MEK-1 and ERK-2 to the nucleus. Using a variety of caveolin-1 deletion mutants, we mapped this in vivo inhibitory activity to caveolin-1 residues 32-95. Peptides derived from this region of caveolin 1 also inhibit the in vitro kinase activity of purified MEK-1 and ERK-2. Thus, we show here that caveolin-1 expression can inhibit signal transduction from the p42/44 MAP kinase cascade both in vitro and in vivo. Taken together with previous data, our results also suggest that a novel form of reciprocal negative regulation exists between p42/44 MAP kinase activation and caveolin-1 protein expression, i.e. up-regulation of caveolin-1 protein expression down-modulates p42/44 MAP kinase activity (this report) and up-regulation of p42/44 MAP kinase activity down-regulates caveolin-1 mRNA and protein expression.

Animals↗
PubMed1998

Crowded little caves: structure and function of caveolae.

Caveolae are small vesicular invaginations of the cell membrane. It is within this organelle that cells perform transcytosis, potocytosis and signal transduction. These "little caves" are composed of a mixture of lipids and proteins unlike those found in the plasma membrane proper. The chief structural proteins of caveolae are caveolins. To date, three caveolins (Cav-1, -2 and -3) with unique tissue distributions have been identified. Caveolins form a scaffold onto which many signalling molecules can assemble, to generate pre-assembled signalling complexes. In addition to concentrating these signal transducers within a distinct region of the plasma membrane, caveolin binding may functionally regulate the activation state of caveolae-associated signalling molecules.

Caveolin 1↗
PubMed1997

Cell-type and tissue-specific expression of caveolin-2. Caveolins 1 and 2 co-localize and form a stable hetero-oligomeric complex in vivo.

Caveolae are microdomains of the plasma membrane that have been implicated in organizing and compartmentalizing signal transducing molecules. Caveolin, a 21-24-kDa integral membrane protein, is a principal structural component of caveolae membrane in vivo. Recently, we and other laboratories have identified a family of caveolin-related proteins; caveolin has been re-termed caveolin-1. Here, we examine the cell-type and tissue-specific expression of caveolin-2. For this purpose, we generated a novel mono-specific monoclonal antibody probe that recognizes only caveolin-2, but not caveolins-1 and -3. A survey of cell and tissue types demonstrates that the caveolin-2 protein is most abundantly expressed in endothelial cells, smooth muscle cells, skeletal myoblasts (L6, BC3H1, C2C12), fibroblasts, and 3T3-L1 cells differentiated to adipocytes. This pattern of caveolin-2 protein expression most closely resembles the cellular distribution of caveolin-1. In line with these observations, co-immunoprecipitation experiments with mono-specific antibodies directed against either caveolin-1 or caveolin-2 directly show that these molecules form a stable hetero-oligomeric complex. The in vivo relevance of this complex was further revealed by dual-labeling studies employing confocal laser scanning fluorescence microscopy. Our results indicate that caveolins 1 and 2 are strictly co-localized within the plasma membrane and other internal cellular membranes. Ultrastructurally, this pattern of caveolin-2 localization corresponds to caveolae membranes as seen by immunoelectron microscopy. Despite this strict co-localization, it appears that regulation of caveolin-2 expression occurs independently of the expression of either caveolin-1 or caveolin-3 as observed using two different model cell systems. Although caveolin-1 expression is down-regulated in response to oncogenic transformation of NIH 3T3 cells, caveolin-2 protein levels remain unchanged. Also, caveolin-2 protein levels remain unchanged during the differentiation of C2C12 cells from myoblasts to myotubes, while caveolin-3 levels are dramatically induced by this process. These results suggest that expression levels of caveolins 1, 2, and 3 can be independently up-regulated or down-regulated in response to a variety of distinct cellular cues.

3T3 Cells↗
PubMed1997

Recombinant expression of caveolin-1 in oncogenically transformed cells abrogates anchorage-independent growth.

Caveolae are plasma membrane-attached vesicular organelles. Caveolin-1, a 21-24-kDa integral membrane protein, is a principal component of caveolae membranes in vivo. Both caveolae and caveolin are most abundantly expressed in terminally differentiated cells: adipocytes, endothelial cells, and muscle cells. Conversely, caveolin-1 mRNA and protein expression are lost or reduced during cell transformation by activated oncogenes such as v-abl and H-ras (G12V); caveolae are absent from these cell lines. However, its remains unknown whether down-regulation of caveolin-1 protein and caveolae organelles contributes to their transformed phenotype. Here, we have expressed caveolin-1 in oncogenically transformed cells under the control of an inducible-expression system. Regulated induction of caveolin-1 expression was monitored by Western blot analysis and immunofluorescence microscopy. Our results indicate that caveolin-1 protein is expressed well using this system and correctly localizes to the plasma membrane. Induction of caveolin-1 expression in v-Abl-transformed and H-Ras (G12V)-transformed NIH 3T3 cells abrogated the anchorage-independent growth of these cells in soft agar and resulted in the de novo formation of caveolae as seen by transmission electron microscopy. Consistent with its antagonism of Ras-mediated cell transformation, caveolin-1 expression dramatically inhibited both Ras/MAPK-mediated and basal transcriptional activation of a mitogen-sensitive promoter. Using an established system to detect apoptotic cell death, it appears that the effects of caveolin-1 may, in part, be attributed to its ability to initiate apoptosis in rapidly dividing cells. In addition, we find that caveolin-1 expression levels are reversibly down-regulated by two distinct oncogenic stimuli. Taken together, our results indicate that down-regulation of caveolin-1 expression and caveolae organelles may be critical to maintaining the transformed phenotype in certain cell populations.

3T3 Cells↗