大鼠肽基甘氨酸α-酰胺化单加氧酶活性及mRNA的组织特异性表达

Tissue specific expression of rat peptidylglycine alpha-amidating monooxygenase activity and mRNA.

作者信息

Braas K M, Stoffers D A, Eipper B A, May V

机构信息

Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.

出版信息

Mol Endocrinol. 1989 Sep;3(9):1387-98. doi: 10.1210/mend-3-9-1387.

DOI:10.1210/mend-3-9-1387

PMID:2575217

Abstract

The tissue specific expression of peptidylglycine alpha-amidating monooxygenase [(PAM) EC 1.14.17.3], an enzyme which catalyzes the formation of amidated bioactive peptides from their glycine-extended precursors, was examined in adult rat. Soluble and membrane-associated PAM enzymatic activities were determined, and the levels and size classes of PAM mRNA were examined by Northern blot analysis. PAM specific activity varied 1000-fold in the tissues examined, with highest levels in heart atrium, pituitary and salivary glands, and hypothalamus. The fraction of total PAM activity that was membrane associated varied from approximately 70% in heart atrium to 10% in neurointermediate pituitary lobe and thyroid gland. Levels of PAM mRNA varied over 300-fold. In the heart atrium, PAM mRNA accounts for more than 0.1% of the mRNA. For many tissues the ratio of total PAM specific activity to PAM mRNA levels was similar; however, PAM activity was higher than expected from mRNA levels in the salivary glands and lower than expected in several tissues, including heart ventricle. Three major size classes of PAM mRNA were identified among the tissues. Use of RNAse H indicated that differences in size were not due to the length of the poly(A) tail. The heart and central nervous system expressed PAM mRNA of the 4.2 kilobase (kb) and 3.8 kb size classes, while the remaining tissues expressed predominantly 3.8 kb and 3.6 kb classes; few tissues contained only one size class of PAM mRNA. The two major forms of PAM mRNA in adult heart atrium differ by the presence or absence of a 315 nucleotide segment in the protein coding region. Using a cDNA probe from within this segment, the 4.2 kb and 3.8 kb size classes of PAM mRNA in the central nervous system appeared to resemble those in the heart atrium. In the remaining tissues, a subset of PAM mRNAs in the 3.8 kb and 3.6 kb size classes hybridized with this probe, suggesting that additional forms of PAM mRNA are present.

摘要

在成年大鼠中检测了肽基甘氨酸α-酰胺化单加氧酶[(PAM)，EC 1.14.17.3]的组织特异性表达，该酶催化从其甘氨酸延伸前体形成酰胺化生物活性肽。测定了可溶性和膜相关的PAM酶活性，并通过Northern印迹分析检测了PAM mRNA的水平和大小类别。在所检测的组织中，PAM比活性变化了1000倍，在心房、垂体、唾液腺和下丘脑中水平最高。膜相关的PAM总活性比例在心房中约为70%，在神经垂体中叶和甲状腺中为10%。PAM mRNA水平变化超过300倍。在心房中，PAM mRNA占mRNA的0.1%以上。对于许多组织，PAM总比活性与PAM mRNA水平的比值相似；然而，唾液腺中的PAM活性高于根据mRNA水平预期的值，而在包括心室在内的几个组织中则低于预期值。在这些组织中鉴定出了三种主要大小类别的PAM mRNA。使用RNA酶H表明大小差异不是由于聚(A)尾的长度。心脏和中枢神经系统表达4.2千碱基(kb)和3.8 kb大小类别的PAM mRNA，而其余组织主要表达3.8 kb和3.6 kb类别；很少有组织只包含一种大小类别的PAM mRNA。成年心房中PAM mRNA的两种主要形式在蛋白质编码区存在或不存在315个核苷酸片段上有所不同。使用该片段内的cDNA探针，中枢神经系统中4.2 kb和3.8 kb大小类别的PAM mRNA似乎与心房中的相似。在其余组织中，3.8 kb和3.6 kb大小类别的PAM mRNA的一个子集与该探针杂交，表明存在其他形式的PAM mRNA。