Anin situatlas of mitochondrial DNA in mammalian tissues reveals high content in stem/progenitor cells

Abstract

<jats:title>ABSTRACT</jats:title><jats:p>Mitochondria regulate ATP production, metabolism and cell death. Alterations in mitochondrial DNA (mtDNA) sequence and copy number are implicated in aging and organ dysfunction in diverse inherited and sporadic diseases. Since most measurements of mtDNA use homogenates of complex tissues, little is known about cell type-specific mtDNA copy number heterogeneity in normal physiology, aging and disease. Thus, the precise cell types whose loss of mitochondrial activity and altered mtDNA copy number that result in organ dysfunction in aging and disease have often not been clarified. Here, we validated an <jats:italic>in situ</jats:italic> hybridization approach to generate a single cell resolution atlas of mtDNA content in mammalian tissues. In hierarchically organized self-renewing tissues, higher levels of mtDNA were observed in stem/proliferative compartments compared to differentiated compartments. Striking zonal patterns of mtDNA levels in the liver reflected the known oxygen tension gradient. In the kidney, proximal and distal tubules had markedly higher mtDNA levels compared to cells within glomeruli and collecting duct epithelial cells. Decreased mtDNA levels were visualized in renal tubules as a function of aging, which was prevented by calorie restriction. We provide a novel approach for quantifying species- and cell type-specific mtDNA copy number and dynamics in any normal or diseased tissue and can be used for monitoring the effects of interventions in animal and human studies.</jats:p>

GSTP1 positive prostatic adenocarcinomas are more common in Black than White men in the United States

“…We analytically validated an automated IHC assay against human GSTP1 protein using well-known cell lines and controls and human prostate cancer tissues(8). Cell line FFPE blocks were prepared and utilized as described(29,30). LNCaP cells were negative and DU145 and PC3 prostate cancer cell lines were positive for GSTP1 protein as expected (2).…”

Visualizing, quantifying, and manipulating mitochondrial DNA in vivo

“…In situ hybridization probes can detect mtDNA and have been used in many applications, including the comparison of mtDNA copy numbers in different tissues (6), visualizing the removal of deleterious mtDNA from the germline after mitochondrial fragmentation (12), and visualization of mtDNA association with particular nucleoids and the resulting genetic autonomy of nucleoids (28).…”

“…5-methylcytosine (5mC) is common in nuclear DNA but is considered to occur less frequently in mtDNA (56). In contrast, N 6 methyldeoxyadenosine (6mA) methylation is reported to occur at 1,300-fold higher levels in mtDNA relative to nuclear DNA and reduces transcription of mtDNA (56). This characteristic methylation signature of mtDNA may play important physiological roles and may also provide an avenue for selective visualization and manipulation of 6mA-methylated mtDNA.…”

“…Mutations of mtDNA can cause a spectrum of mitochondrial diseases where the clinical expression depends on the specific mutation and the degree of heteroplasmy (the proportion of mutated mtDNA in a single cell) between wild-type and mutant mtDNA (3,4). The copy number of mtDNA is controlled by poorly understood mechanisms, but varies between tissues, during ageing, and in cancer (5)(6)(7). The mtDNA within mitochondria is present within nucleoids (8).…”

“…Direct visualisation of mtDNA in vivo can thus offer further mechanistic insight. Visualization of the mtDNA copy number has revealed that mtDNA increases its population during S-phase in the cell cycle (17), that mtDNA copy number differs between tissues and can decline during ageing (6), and that mtDNA copy number is reduced in some cancers such as glioma (18). Visualization of mtDNA in yeast has shown that segregation of mtDNA during cell division preserves the density of mtDNA in daughter cells, in part via the semi-regular spacing of nucleoids within mitochondria (19,20).…”