Protein degradation during terminal cytodifferentiation. Studies on mammary gland in organ culture

1. In mammary gland explants subjected to experimental manipulation, average rates (during 24 h periods) of degradation of fatty acid synthase, casein and cytosol-fraction proteins were measured by a double-isotope method. Rates of degradation of fatty acid synthase were also computed from measurements of changing enzyme amount and rate of synthesis. 2. During the period of most rapid enzyme accumulation there is a transient decrease in the computed rate of degradation of fatty acid synthase. Removal of hormones produces a rapid increase in the computed rate of degradation of the enzyme. 3. The average rate of degradation of fatty acid synthase measured by the double-isotope method is low in the presence of hormones, and increases on hormone removal. This increase in degradation rate is inhibited by adrenaline and further blocked by insulin. NH4Cl (10 mM) also partially inhibits the increase in protein degradation on hormone removal. 4. The pattern of changes in the average rate of degradation of cytosol-fraction proteins is similar to that for fatty acid synthase alone. There is no relationship between subunit molecular weight and rate of degradation under all experimental conditions. 5. Isotope ratios for resolved cytosol protein mixtures are transformed logarithmically to make the standard deviations an estimate of heterogeneity of degradation rates. By this analysis, in some conditions there appears to be significant measureable heterogeneity of degradation rates. 6. Little degradation of casein is measured in the presence of hormones, but a marked increase in the rate of degradation can be measured when hormones are removed. Whereas at 24-48h NH4Cl (10 mM) has little effect on this enhanced rate of degradation, at 48-72h it causes a large decrease in degradation rate. 7. Results are discussed in terms of a two-component degradation system in mammary gland explants.

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Regulation of lipogenic capacity in lactating rats

Biochemical Journal ◽

10.1042/bj2080611 ◽

1982 ◽

Vol 208 (3) ◽

pp. 611-618 ◽

Cited By ~ 12

Author(s):

M R Grigor ◽

A Geursen ◽

M J Sneyd ◽

S M Warren

Keyword(s):

Mammary Gland ◽

Fatty Acid ◽

Malic Enzyme ◽

Fatty Acid Synthase ◽

Specific Activity ◽

Mammary Glands ◽

Lipogenic Enzymes ◽

Pregnancy And Lactation ◽

Specific Activities

1. The rate of mammary-gland lipogenesis measured in vivo from 3H2O was suppressed after decreasing the milk demand by decreasing the number of pups from ten to two or three, as well as by giving diets containing lipid [Grigor & Warren (1980) Biochem. J. 188, 61-65]. 2. The specific activities of the lipogenic enzymes fatty acid synthase, glucose 6-phosphate dehydrogenase and ‘malic’ enzyme increased between 6- and 10-fold in the mammary gland and between 2- and 3-fold in the livers during the first 10 days of lactation. The increases in specific activity coupled with the doubling of liver mass which occurred during pregnancy and lactation resulted in considerable differences in total liver activities when compared with virgin animals. 3. Although consumption of a diet containing 20% peanut oil suppressed the activities of the three lipogenic enzymes in the livers, only the ‘malic’ enzyme was affected in the mammary glands. 4. In contrast, decreased milk demand did not affect the specific activities of any of the liver enzymes, whereas it resulted in suppression of all three lipogenic enzymes of the mammary glands. There was no effect on either the cytoplasmic malate dehydrogenase or the lactate dehydrogenase of the mammary gland. 5. In all the experiments performed, the activity of the fatty acid synthase correlated with the amount of material precipitated by the rabbit antibody raised against rat fatty acid synthase.

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Rat mammary-gland fatty acid synthase. A simple purification procedure and stoicheiometry of CoA ester binding

Biochemical Journal ◽

10.1042/bj2030045 ◽

1982 ◽

Vol 203 (1) ◽

pp. 45-50 ◽

Cited By ~ 4

Author(s):

P M Ahmad ◽

D S Feltman ◽

F Ahmad

Keyword(s):

Mammary Gland ◽

Fatty Acid ◽

Fatty Acid Synthase ◽

Specific Activity ◽

Simple Procedure ◽

Polyacrylamide Gel Electrophoresis ◽

Sodium Dodecyl ◽

Performic Acid ◽

Mammary Tissue ◽

Rat Mammary Gland

A simple procedure was devised which allows purification of rat lactating-mammary-gland fatty acid synthase to a high degree of purity, with recoveries of activity exceeding 50%. Over 50 mg of enzyme was isolated from 60 g of mammary tissue. The specific activity of the purified enzyme was about 2.5 mumol of NADPH oxidized/min per mg of protein at 37 degrees. The enzyme appeared homogeneous by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and by immunodiffusion analysis. Each mol (Mr 480 000) of the enzyme bound 3 mol of acetyl and 3-4 mol of malonyl groups when the binding experiments were performed at 0 degrees for 30 s. The presence of NADPH did not influence the binding stoicheiometry for these acyl-CoA derivatives. Approx. 2 mol of taurine was found per mol of the performic acid-oxidized enzyme, suggesting that there were 2 mol of 4′-phosphopantetheine in the native enzyme. Rat mammary-gland fatty acid synthase required free CoA for activity.

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Association of cytosolic lipids with fatty acid synthase from lactating mammary gland

International Journal of Biochemistry ◽

10.1016/0020-711x(93)90661-w ◽

1993 ◽

Vol 25 (4) ◽

pp. 533-543 ◽

Cited By ~ 7

Author(s):

Brigitte H. Keon ◽

Debabrota Ghosal ◽

Thomas W. Keenan

Keyword(s):

Mammary Gland ◽

Fatty Acid ◽

Fatty Acid Synthase ◽

Lactating Mammary Gland

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Repurposing combination therapy of Voacamine with Vincristine for down regulation of HIF-1α/FASN co-axis and PHD2 activation in ER+ mammary neoplasia

10.21203/rs.3.rs-61541/v1 ◽

2020 ◽

Author(s):

Lakhveer Singh ◽

Manjari Singh ◽

Dinesh Kumar ◽

Mohd. Nazam Ansari ◽

Abdulaziz S. Saeedan ◽

...

Keyword(s):

Mammary Gland ◽

Fatty Acid ◽

Combination Therapy ◽

Fatty Acid Synthase ◽

Hypoxia Inducible Factor ◽

Prolyl Hydroxylase ◽

Over Expression ◽

Hypoxia Inducible Factor 1Α ◽

Gland Carcinoma

Abstract Background The current study was attempted to inquest the role of combination therapy of Voacamine and Vincristine for the prevention of mammary gland carcinoma through prolyl hydroxylase-2 activation. The prolyl hydroxylase‐2 activation leads the downregulation of hypoxia‐inducible factor‐1α and fatty acid synthase. Over expression of hypoxia inducible factor-1α and fatty acid synthase is previously reported in solid tumor of mammary gland. Methods After screening a battery of natural compounds which were similar to vincristine, vocamine was selected as a possible prolyl hydroxylase-2 activator and justify its activity using 7, 12-Dimethylbenz[a]anthracene induced rat model. The combination therapy was evaluated for cardiac toxicity using hemodynamic profile. The angiogenic markers were evaluated using carmine staining. Monotherapy and combination therapy were also evaluated for liver and kidney toxicity through haematoxylin and eosin staining. The combination therapy also delineated the markers of oxidative stress favorably. Afterwards, the disruption of fatty acids was evaluated using gas chromatography. Results The immunoblotting analysis validated that combination therapy has a potential to switch on the prolyl hydroxylase-2 activity and thus initiate proteolytic degradation of hypoxia‐inducible factor‐1α and its consequence effects. The combination therapy also stimulated programmed cell death (apoptosis) in rapidly dividing cancer cells. Conclusion The present study explores the role of voacamine in activation of prolyl hydroxylase-2 which can decrease over expression of hypoxia‐inducible factor‐1α and fatty acid synthase in cells of mammary gland carcinoma.

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Repurposing Combination Therapy of Voacamine with Vincristine for Down Regulation of HIF-1α/FASN Co-Axis and PHD2 Activation in ER+ Mammary Neoplasia

10.21203/rs.3.rs-34012/v1 ◽

2020 ◽

Author(s):

Lakhveer Singh ◽

Manjari Singh ◽

Mohd. Nazam Ansari ◽

Abdulaziz S. Saeedan ◽

Gaurav Kaithwas

Keyword(s):

Mammary Gland ◽

Fatty Acid ◽

Combination Therapy ◽

Fatty Acid Synthase ◽

Hypoxia Inducible Factor ◽

Prolyl Hydroxylase ◽

Over Expression ◽

Hypoxia Inducible Factor 1Α ◽

Gland Carcinoma

Abstract Background: The current study was attempted to inquest the role of combination therapy of Voacamine and Vincristine for the prevention of mammary gland carcinoma through prolyl hydroxylase‐2 activation. The prolyl hydroxylase‐2 activation leads the downregulation of hypoxia‐inducible factor‐1α and fatty acid synthase. Over expression of hypoxia inducible factor-1α and fatty acid synthase is previously reported in solid tumor of mammary gland. Methods: After screening a battery of natural compounds which were similar to vincristine, vocamine was selected as a possible prolyl hydroxylase‐2 activator and justify its activity using 7, 12-Dimethylbenz[a]anthracene induced rat model. The combination therapy was evaluated for cardiac toxicity using hemodynamic profile. The angiogenic markers were evaluated using carmine staining. Monotherapy and combination therapy were also evaluated for liver and kidney toxicity through haematoxylin and eosin staining. The combination therapy also delineated the markers of oxidative stress favorably. Afterwards, the disruption of fatty acids was evaluated using gas chromatography. Results: The immunoblotting analysis validated that combination therapy has a potential to switch on the prolyl hydroxylase‐2 activity and thus initiate proteolytic degradation of hypoxia‐inducible factor‐1α and its consequence effects. The combination therapy also stimulated programmed cell death (apoptosis) in rapidly dividing cancer cells.Conclusion: The present study explores the role of voacamine in activation of prolyl hydroxylase‐2 which can decrease over expression of hypoxia‐inducible factor‐1α and fatty acid synthase in cells of mammary gland carcinoma.

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Studies on acetyl-CoA carboxylase and fatty acid synthase from rat mammary gland and mammary tumours

Biochemical Journal ◽

10.1042/bj2080443 ◽

1982 ◽

Vol 208 (2) ◽

pp. 443-452 ◽

Cited By ~ 2

Author(s):

P M Ahmad ◽

D S Feltman ◽

F Ahmad

Keyword(s):

Mammary Gland ◽

Fatty Acid ◽

Fatty Acid Synthase ◽

Mammary Tissue ◽

Acetyl Coa Carboxylase ◽

Lipogenic Enzymes ◽

Acetyl Coa ◽

Rat Mammary Gland ◽

Mammary Gland Differentiation ◽

Mammary Adenocarcinomas

The activities of two lipogenic enzymes, acetyl-CoA carboxylase and fatty acid synthase, were determined in two transplantable mammary adenocarcinomas (13762 and R3230AC) carried by non-pregnant, pregnant and lactating rats, and in mammary tissue of control animals (non-tumour-carrying) of comparable physiological states. During mammary-gland differentiation of control or tumour-carrying animals, the activities of acetyl-CoA carboxylase and fatty acid synthase in the lactating gland increased by about 40-50-fold over the values found in non-pregnant animals. On the other hand, in tumours carried by lactating dams there were only modest increases (1.5-2-fold) in acetyl-CoA carboxylase and fatty acid synthase compared with the neoplasms carried by non-pregnant animals. On the basis of the Km values for different substrates and immunodiffusion and immunotitration data, the fatty acid synthase of neoplastic tissues appeared to be indistinguishable from the control mammary-gland enzyme. However, a comparison of the immunotitration and immunodiffusion experiments indicated that the mammary-gland acetyl-CoA carboxylase might differ from the enzyme present in mammary neoplasms.

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