Synchronized transcriptional gene expression of H⁺‐ATP synthase subunits in different tissues of Fischer 344 rats of different ages

Abstract

Little is known about the relationship between the stoichiometry of polypeptides of multisubunit enzyme complexes and the absolute amount of each transcript of the complexes in mammalian tissues. Here we showed that the absolute amounts of the transcripts of most subunits of rat H 1 -ATP synthase examined greatly differed in the different tissues, showing the following hierarchy of tissue-specificity: heart . kidney . brain < liver. However, surprisingly, there was no difference in the expression pattern of these in terms of the molar ratio of each transcript, indicating a nearly similar stoichiometric expression pattern irrespective of tissue or age of the rat. Therefore, the present finding clearly indicates that most of the transcripts of the 16 subunits of rat H 1 -ATP synthase were concertedly and synchronously expressed, having a constant expression pattern of the transcripts, irrespective of tissue or age of the rats. This is the first report of the absolute amounts of the transcripts of this multisubunit enzyme.Keywords: H 1 -ATP synthase; expression pattern; transcript; stoichiometry; synchronization.The mechanism of the transcriptional regulation of the genes involved in the biosynthesis and assembly of multisubunit mitochondrial enzyme complexes, e.g. Complex I, Complex III, Complex IV and H 1 -ATP synthase, is still obscure [1±4]. Of particular interest is the biogenesis of H 1 -ATP synthase, the world's smallest rotary motor [5±9], the key enzyme in oxidative phosphorylation and the one responsible for the production of most of the ATP in mammalian organisms. Mammalian H 1 -ATP synthase is a macromolecular supercomplex composed of 16 subunits [10±14] with a constant stoichiometry, 6 of which construct the catalytic site of ATP synthesis called F 1 (subunits a 3 , b 3 , g 1 , d 1 , and : 1 , and the loosely attached ATPase inhibitor protein IF1 1 ) [15,16] and 10 of which construct the energy-transduction part called F 0 (subunits a 1 , b 2 , c 10 , d 1 , e 2 , f unknown , g unknown , OSCP 1 , F6 2 , and A6L 1 ) [14,17±18a]. Subunit a and A6L are encoded by the mitochondrial genome, and all of the other subunits are separately encoded by nuclear chromosomes; however, the relationship between the stoichiometry of polypeptides of the complexes and the absolute amount of each transcript of this multisubunit enzyme complex in mammalian tissues is still entirely unknown.In the present work, we determined the absolute amount of nine species of mRNAs for eight nucleus-encoded subunits of H 1 -ATP synthase in the brain, liver, heart, and kidney tissues of 2-week-old to 90-week-old male Fischer 344 rats by using highly purified poly(A) 1 RNA and the mRNAs synthesized by an in vitro transcriptional system. M A T E R I A L S A N D M E T H O D S AnimalsMale Fischer 344 rats were obtained from Charles River Japan Inc (Atsugi, Japan), and were genetically identical because of approximately 300 matings between sister and brother. Purification of Poly(A) 1 RNAThe rats were decapitated, and tissues of brain, liver, h...

Functional Disorders of the Oxidative Phosphorylation System in the Heart Mitochondria of Mice with Juvenile Visceral Steatosis.

“…36) As shown in this present study, the expression levels of a subunit and CF 6 of H ϩ -ATP synthase in the heart mitochondria from control and JVS mice were almost the same (Fig. 1).…”

Identification of the Up- and Down-Regulated Genes in the Heart of Juvenile Visceral Steatosis Mice

“…For light microscopy, hearts were fixed in 10% phosphate-buffer (pH 7.4), dehydrated in a graded series of ethanol, and embedded in paraffin, after 12) and 9-week-old control and JVS mice by the guanidinium thiocyanate method as described previously. 13) Poly(A) ϩ RNA was purified from total RNA from 9-weekold control and JVS mice by Oligotex-dT30 ͗Super͘ mRNA Purification Kit (TaKaRa).…”

“…Dot Blot Analysis Dot blot analysis using 100 ng of poly(A) ϩ RNA from 9-week-old control and JVS mice and quantitative determination of 5 subunits of H ϩ -ATP synthase, c(P1), c(P2), e, ATPase inhibitor protein IF1, and b was carried out as described previously. 13) RESULTS AND DISCUSSION Morphological Study Hearts weighed 0.06Ϯ0.02 g in 2-week-old JVS mice and 0.03Ϯ0.01 g in 2-week-old control mice (the values are the meanϮS.D. for 8 and 20 experiments with JVS and control mice, respectively; pϽ0.00001, JVS versus control mice).…”