Localization of nuclear subunits of cyclic AMP-dependent protein kinase by the immunocolloidal gold method.
Localization of nuclear subunits of cyclic AMP-dependent protein kinase by the immunocolloidal gold method.
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通过免疫固定金方法的环状AMP依赖性蛋白激酶的核亚基定位。
DOI：
10.1083/jcb.101.3.965
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发表时间：
1985-09
影响因子：
7.8
通讯作者：
JUNGMANN, RA
中科院分区：
文献类型：
--
作者：
KUETTEL, MR;SQUINTO, SP;KWASTWELFELD, J;SCHWOCH, G;SCHWEPPE, JS;JUNGMANN, RA
介绍了一种免疫胶体金电子显微镜方法，可以对cAMP依赖的蛋白激酶的调节亚基RI和RII以及催化亚基C进行超微结构定位和定量。利用包埋后的间接免疫金标记法，利用特定的抗血清，催化和调节亚基被定位在核的电子致密区和细胞质区域，非特异性染色最少。抗原域定位于大鼠肝细胞、卵巢颗粒细胞、精原细胞以及培养的H4IIE肝癌细胞的异染色质、核仁、染色质间颗粒和内质网中。核抗原相对染色密度的形态计量学定量表明，在不同的生理条件下，单位面积上的亚基数量有明显的变化。例如，大鼠肝部分切除后，核内RI和C亚基的染色密度在术后16h显著增加。胰升糖素处理的大鼠只增加了核催化亚基的染色密度。二丁酰cAMP处理H4IIE肝癌细胞后，cAMP依赖蛋白激酶三个亚基的核染色密度均显著增加。这些研究表明，针对cAMP依赖的蛋白激酶亚基的特异性抗血清可以与免疫金电子显微镜结合使用，以确定亚基的超微结构位置，并提供其相对细胞密度的半定量估计。
An immunocolloidal gold electron microscopy method is described allowing the ultrastructural localization and quantitation of the regulatory subunits RI and RII and the catalytic subunit C of cAMP- dependent protein kinase. Using a postembedding indirect immunogold labeling procedure that employs specific antisera, the catalytic and regulatory subunits were localized in electron-dense regions of the nucleus and in cytoplasmic areas with a minimum of nonspecific staining. Antigenic domains were localized in regions of the heterochromatin, nucleolus, interchromatin granules, and in the endoplasmic reticulum of different cell types, such as rat hepatocytes, ovarian granulosa cells, and spermatogonia, as well as cultured H4IIE hepatoma cells. Morphometric quantitation of the relative staining density of nuclear antigens indicated a marked modulation of the number of subunits per unit area under various physiologic conditions. For instance, following partial hepatectomy in rats, the staining density of the nuclear RI and C subunits was markedly increased 16 h after surgery. Glucagon treatment of rats increased the staining density of only the nuclear catalytic subunit. Dibutyryl cAMP treatment of H4IIE hepatoma cells led to a marked increase in the nuclear staining density of all three subunits of cAMP-dependent protein kinase. These studies demonstrate that specific antisera against cAMP-dependent protein kinase subunits may be used in combination with immunogold electron microscopy to identify the ultrastructural location of the subunits and to provide a semi-quantitative estimate of their relative cellular density.
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影响因子：
3.2
作者：
BENDAYAN, M
通讯作者：
BENDAYAN, M
影响因子：
4.8
作者：
HUNZICKERDUNN, M;JUNGMANN, RA;WEST, DE
通讯作者：
WEST, DE
DOI：
10.1073/pnas.79.17.5137
发表时间：
1982-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
LAMERS, WH;HANSON, RW;MEISNER, HM
通讯作者：
MEISNER, HM
影响因子：
3.2
作者：
BENDAYAN, M;ZOLLINGER, M
通讯作者：
ZOLLINGER, M
影响因子：
7.8
作者：
BYUS, CV;FLETCHER, WH
通讯作者：
FLETCHER, WH