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B B Mukherjee

Publications and source records attributed to B B Mukherjee.

At least 37 records · Page 2Linked to original sources
PubMed1988

Dibutyryl cAMP inhibits expression of transformation-related properties in Kirsten murine sarcoma virus transformed Balb/c-3T3 cells despite continued presence of p21v-Ki-ras.

We studied the effects of simultaneous treatment with 0.1 mM N6, O2'-dibutyryl cAMP (dbcAMP) and 1 mM theophylline on several transformation-specific properties and on levels of the Kirsten murine sarcoma virus (Ki-MSV) transforming gene product p21v-Ki-ras, in a Ki-MSV-transformed mouse cell line (Balb/c-3T3, clone A31; KA31). The rate of logarithmic growth, cell motility, and final saturation density were reduced in dbcAMP-treated KA31 cultures. Capabilities for anchorage-independent growth were reduced in treated cells, to levels similar to those observed for the untransformed parental A31 cell line. Treatment with dbcAMP had no observable effect on the binding of 125I-labeled epidermal growth factor and did not alter fluorescence staining patterns for actin microfilaments and fibronectin which, although characteristic of normal cells, were also present in KA31 cells. Changes induced by dbcAMP were readily reversible, except for loss of anchorage-independent growth. However, this property was also reversible, provided removal of dbcAMP occurred 48 h prior to inoculation into soft agar medium. Immunoprecipitation with a monoclonal antibody directed against the protein p21v-Ki-ras (Y13-259) revealed the continued presence of this protein in dbcAMP-treated KA31 cells. We, therefore, conclude that cAMP mediates the inhibition of growth-related transformation-specific properties either by acting at steps subsequent to the expression of p21v-Ki-ras or on a pathway independent of p21ras function.

Actins↗
PubMed1987

Epidermal and transforming growth factors modulate secretion of a 69 kDa phosphoprotein in normal rat kidney fibroblasts.

Our study shows that the secretion of a major non-glycosylated, phosphoprotein of 69 kDa (pp69) is a specific marker for non-transformed NRK-49F cells. Treatment of NRK-49F cells with EGF alone or with different combinations of EGF plus TGF-beta modulates the secretion of pp69, suggesting its relationship with cellular proliferation. Antibody raised against pp69 recognizes, in addition to pp69, another major phospho-protein of 62 kDa (pp62) secreted by RR1022 and spontaneously transformed NRK-49F cells. Immunoprecipitation of total cell lysates from both NRK-49F and RR1022 cells with anti-pp69 antibody detected only pp69. These observations suggest a precursor-product relationship between pp69 secreted by non-transformed NRK-49F cells and pp62 secreted by transformed cells.

Animals↗
PubMed1986

Ability of oncogenically transformed cells to grow without anchorage correlates with phosphorylation of a group of cell surface membrane proteins.

Anchorage-independent growth in vitro is strongly correlated with cellular malignancy in vivo and it has been shown that retinoic acid (RA; a vitamin A analog) inhibits anchorage-independent growth of a wide variety of oncogenically transformed cells (RA-sensitive cells). We report here that decreased or lack of phosphorylation of a group of low molecular weight (20-30 kD) cell surface membrane proteins, particularly one of Mr 28 kD, correlates strongly with RA-induced loss of anchorage-independent growth of RA-sensitive cells. Our studies also show that this group of proteins are not phosphorylated in non-transformed cells which do not grow in an anchorage-independent manner. Analysis of [35S]methionine-labeled proteins revealed that these polypeptides are present in both RA-treated and untreated cell surface membranes. This suggests that modulation of phosphorylation rather than lack of synthesis of these proteins is correlated with anchorage regulation of cells. V8 protease mapping of the 28 kD phosphoprotein from transformed cells, irrespective of their origin or of transforming agents, revealed complete fragment homology. Furthermore, the 28 kD phosphoprotein was found to be phosphorylated exclusively at threonine residues. The data obtained from this study suggest that the ability of cells to grow without anchorage is correlated with the phosphorylation of a group of cell surface membrane proteins and RA inhibits anchorage-independent growth by interfering with the phosphorylation rather than synthesis of these proteins.

Animals↗
PubMed1986

Altered biochemical properties of actin in normal skin fibroblasts from individuals predisposed to dominantly inherited cancers.

The data presented here show that normal skin fibroblasts from individuals with dominantly inherited retinoblastoma, polyposis coli, and nevoid basal cell carcinoma (predisposed cells), grown in the presence of [35S]methionine, contain more than 2.5-fold [35S]methionine-labeled actin as compared to normal fibroblasts from individuals without a prior history of predisposition to cancer (normal cells). The rate of incorporation of [35S]methionine into actin in predisposed cells is rapid and is not correlated with an increase in the total protein and actin contents of the cells, in the intracellular pool size of [35S]methionine, or in the synthesis of beta-actin-specific mRNA, as compared to normal cells. However, the half-life of actin in predisposed cells is less than 5 h, as compared to at least 48 h for normal cells. The significantly reduced half-life of actin and an increased incorporation of [35S]methionine specifically into actin in all predisposed cells studied may represent an inherited biochemical defect which leads to cytoskeletal disorganization previously observed in these cells. It can be speculated that the altered properties of actin in predisposed cells may be caused by the same genetic lesion(s) which is responsible for the induction of dominantly inherited cancers.

Actins↗
PubMed1985

Lack of correlation between loss of anchorage-independent growth and levels of transformation-specific p53 protein in retinoic acid-treated F9 embryonal carcinoma cells.

It has been shown that differentiated derivatives of retinoic acid (RA)-treated F9 embryonal carcinoma cells become non-malignant. In the present study it is asked whether this loss of malignancy is due to cellular differentiation. Because the ability of cells to grow in suspension correlates with in vivo tumorigenicity, we determined the time course of the loss of this property, after RA treatment, with relation to the differentiation to parietal endoderm and the acquisition of normalcy in several common transformation-specific properties of F9 cells. Our results show that pretreatment with RA for 24 h caused 80% inhibition of anchorage-independent growth in F9 cells, and this inhibition reached its highest level (98%) after pretreatment with RA for 48 h and longer. However, all other observed transformation-related properties, and the levels of plasminogen activator (marker for parietal endoderm) remained unaltered at this early post-treatment stage. These observations suggest that the loss of malignancy is a relatively early event in the biochemical pathways involved in the RA-induced differentiation of F9 cells. Furthermore, our data show that the presence of elevated levels of p53 alone may not be sufficient to maintain the anchorage-independent growth and the rapid proliferation of F9 cells.

Actins↗
PubMed1985

Altered processing of a major secreted phosphoprotein correlates with tumorigenicity in Rous sarcoma virus-transformed mammalian cells.

Anchorage-independent growth is highly correlated with neoplastic growth in vivo, and the retinoids (vitamin A and its analogs) inhibit this property in a wide variety of oncogenically transformed cells. We report here that retinoic acid-treated Rous sarcoma virus-transformed rat (RR1022) and vole (SR-1T) cells, which show reversible loss of anchorage-independent growth and assume nontransformed morphology, secrete a major 69-kilodalton phosphoprotein (pp69) instead of the 62-kilodalton phosphoprotein (pp62) secreted by their untreated counterparts. As determined by V8 protease mapping and by two-dimensional electrophoretic analysis, this 69-kilodalton polypeptide was indistinguishable from the pp69 released by nontransformed normal rat kidney cells. Neither retinoic acid-treated RR1022 cells nor normal rat kidney cells secreted pp62, and retinoic acid treatment did not have any significant effect on the synthesis, subcellular localization, or phosphokinase activity of pp60src. Furthermore, treatment with retinoic acid did not alter the synthesis of the transformation-specific 53-kilodalton phosphoprotein (p53) and secretion of the transforming growth factors in RR1022 cells. Our studies showed that there is a clear correlation between the release of pp69 or pp62 and the ability of cells to grow in vitro with or without anchorage. This may provide an important clue for elucidating specific biochemical events involved in anchorage regulation of growth.

Animals↗
PubMed1983

Restoration of anchorage regulation in transformed cells by retinoic acid (RA) is independent of the presence of cytoplasmic RA-binding proteins.

In an attempt to analyse the cause-effect relationship between anchorage-independent growth (a property which correlates best with in vivo tumorigenicity) and a set of other common transformation-related properties, the effect of retinoic acid (RA) treatment on six unrelated transformed cell lines (including DNA tumor virus, retrovirus, and spontaneously transformed cells) were studied. The data show that the changes in morphology and cellular orientation in culture, loss of cell surface fibronectin, disruption of actin microfilaments, increased hexose uptake, loss of density-dependent growth, and decreased binding of EGF, properties which are often associated with oncogenic transformation of cells, are dissociable from one another and from anchorage-independent growth. RA appears to interfere with anchorage-independent growth of all the retrovirus and spontaneously transformed cell lines (responsive cells) that we examined; however, such treatment failed to inhibit anchorage-independent growth in both of the DNA tumor virus-transformed cell lines (non-responsive cells) that we used in the present study. The presence of RA-binding proteins in both responsive and non-responsive cells suggests that the mechanism of RA action for the inhibition of anchorage-independent growth in transformed cells may be independent of the presence of such cytoplasmic proteins. Finally, the present study clearly indicates that the use of RA treatment, like partial transformation mutants of oncogenic viruses, can be a novel approach in analysing the general mechanism by which transformed cells grow without anchorage.

Actins↗
PubMed1982

Effects of 12-O-tetradecanoylphorbol-13-acetate on fibroblasts from individuals genetically predisposed to cancer.

The purpose of the present study was to determine whether skin fibroblasts from individuals, either with an inherited predisposition to cancer or with genetic disorders usually associated with a high risk of cancer, can be oncogenically transformed in vitro by a tumor promoter alone. The effects of chronic and limited applications of 12-O-tetradecanoylphorbol-13-acetate (TPA) on several properties that are associated with transformation were examined using skin fibroblasts from individuals with polyposis coli, a familial cancer syndrome, xeroderma pigmentosum, Fanconi's anemia, and trisomy 21. The results of this study show that TPA treatment induces similar changes on cellular morphology, growth rate, saturation density, epidermal growth factor binding, and cytoskeleton in fibroblasts from both normal and genetically predisposed individuals. None of these cell lines, however, acquired anchorage-independent growth or unlimited growth potential in culture after chronic application of TPA. These observations suggest clearly that skin fibroblasts from individuals with either a genetic predisposition to or a high risk of cancer may not exist in a preneoplastic or "initiated" state susceptible to oncogenic transformation by TPA alone and that the mechanism of genetically determined cancer induction may be different from that of chemical carcinogenesis.

Cell Adhesion↗
PubMed1981

Glycoprotein biosynthesis in animal cells grown in suspension culture. Assembly of lipid-linked saccharides and formation of protein-bound 'high-mannose' oligosaccharides.

Glycoprotein biosynthesis was studied with mouse L-cells grown in suspension culture. Glucose-deprived cells incorporated [3H]mannose into 'high-mannose' protein-bound oligosaccharides and a few relatively high-molecular-weight lipid-linked oligosaccharides. The latter were retained by DEAE-cellulose and turned over quite slowly during pulse--chase experiments. Increased heterogeneity in size of lipid-linked oligosaccharides developed during prolonged glucose deprivation. Sequential elongation of lipid-linked oligosaccharides was also observed, and conditions that prevented the assembly of the higher lipid-linked oligosaccharides also prevented the formation of the larger protein-bound 'high-mannose' oligosaccharides. In parallel experiments, [3H]mannose was incorporated into a total polyribosome fraction, suggesting that mannose residues were transferred co-translationally to nascent protein. Membrane preparations from these cells catalysed the assembly from UDP-N-acetyl-D-[6-3H]glucosamine and GDP-D-[U-14C]mannose of polyisoprenyl diphosphate derivatives whose oligosaccharide moieties were heterogeneous in size. Elongation of the N-acetyl-D-[6-3H]glucosamine-initiated glycolipids with mannose residues produced several higher lipid-linked oligosaccharides similar to those seen during glucose deprivation in vivo. Glucosylation of these mannose-containing oligosaccharides from UDP-D-[6-3H]glucose was restricted to those of a relatively high molecular weight. Protein-bound saccharides formed in vitro were mainly smaller in size than those assembled on the lipid acceptors. These results support the involvement of lipid-linked saccharides in the synthesis of asparagine-linked glycoproteins, but show both in vivo and in vitro that protein-bound 'high-mannose' oligosaccharide formation can occur independently of higher lipid-linked oligosaccharide synthesis.

Animals↗
PubMed1980

Characterization of X-tropic endogenous retrovirus-specific RNA uninfected tissues and lymphosarcoma of BALB/c mice.

Analysis of endogenous X-tropic BALB/c retrovirus-specific RNAs in a BALB/c mouse tissue containing mostly non-driving cells (liver), in two normal tissues having significant proportions of dividing cells (20-h regenerating liver and 12-day embryo) and in cells of a lymphosarcoma of BALB/c mice were carried out by determining the extent to which the RNAs from these tissues hybridized to the virus 3H-cDNA probe and by their relative sedimentation values in a sucrose gradient. RNAs from 20-h regenerating liver, 12-day embryo and lymphosarcoma, each containing a significant proportion of proliferating cells, showed 8 to 21 % higher hybridization values than normal liver RNA. Differences in the exact size classes of virus-specific RNAs and, in their relative proportions were found to exist in the four different tissue types examined and no correlation between a specific RNA size-profile and the proliferating activity of a tissue could be detected.

Animals↗
PubMed1980

Restriction of the rat-specific RNS sequences of Ki-MSV to the nucleus of dibutyryl cyclic AMP-treated K-A31 cells.

The treatment of oncogenically transformed cells in culture, with dibutyryl cyclic AMP (cAMP) has, in many cases, resulted in a general phenotypic change towards the normal state. A virus-specific gene product(s) is responsible for the transformation of cells by sarcoma viruses and it has been suggested that the src gene product may act through the alteration of cAMP levels. With these premises we have studied the effects of dibutyryl cAMP on cell growth and virus genome expression in a Kirsten sarcoma virus-transformed mouse cell line. Our results suggest that certain virus-specific RNA sequences are restricted to the nucleus of these cells after several days of growth in medium containing dibutyryl cAMP and that these sequences appear to be those coding for the sarcoma information.

Animals↗
PubMed1979

In vivo induction of endogenous retroviruses in BALB/c mouse hepatocytes by successive treatments with carbon tetrachloride and bromodeoxyuridine.

The effects of bromodeoxyuridine (BrdUrd) incorporation into hepatocytes of carbon tetrachloride (CCl4)-induced regenerating liver of BALB/c mice on the induction of endogenous retroviruses was examined. From the nucliec acid hybridization studies, the maximum levels of hybridization were obtained for both N- and X-tropic BALB/c endogenous retrovirus specific [3H]-cDNAs with liver RNA from animals receiving BrdUrd at 40 and 44 h post-CCl4 treatment, and killed on the fourth day following BrdUrd injection. Media from NIH-3T3 (Swiss mouse) and mink cell cultures, infected with liver homogenates from animals treated as above, gave significant levels of reverse transcriptase activity. The observations made in the present study show that BrdUrd incorporation into cell DNA can cause induction of both N- and X-tropic endogenous retroviruses in BALB/c mouse hepatocytes in vivo, and such induction is probably a transient event.

Animals↗
PubMed1979

Selective transcription of genomic sequences common to both N- and X-tropic endogenous retroviruses in BALB/c mouse tissues.

Total RNAs from four BALB/c mouse tissues, containing mostly non-dividing cells (liver and kidney) or variable proportions of dividing cells (uterus and embryo) were analysed for sequences complementary to 3H-DNA transcripts synthesized from BALB/c endogenous. N- and X-tropic retroviruses. Extensive transcription of virogene information was detected in the tissues examined, but such transcription was found to be mostly limited to the homologous regions of the two virus genomes. No additivity of hybridization values could be detected when RNAs from two different tissues were mixed, which suggests that BALB/c mouse liver, kidney, uterus and embryo transcribe a common set of nucleic acid sequences of the homologous regions of the N- and X-tropic viral genomes, in addition to other sequences of the same region that are specific for indiviual tissues.

Animals↗