Dataset Information

Phosphorylation and acetylation of mitochondrial transcription factor A promote transcription processivity without compromising initiation or DNA compaction.

ABSTRACT: Mitochondrial transcription factor A (TFAM) plays important roles in mitochondrial DNA compaction, transcription initiation, and in the regulation of processes like transcription and replication processivity. It is possible that TFAM is locally regulated within the mitochondrial matrix via such mechanisms as phosphorylation by protein kinase A and nonenzymatic acetylation by acetyl-CoA. Here, we demonstrate that DNA-bound TFAM is less susceptible to these modifications. We confirmed using EMSAs that phosphorylated or acetylated TFAM compacted circular double-stranded DNA just as well as unmodified TFAM and provide an in-depth analysis of acetylated sites on TFAM. We show that both modifications of TFAM increase the processivity of mitochondrial RNA polymerase during transcription through TFAM-imposed barriers on DNA, but that TFAM bearing either modification retains its full activity in transcription initiation. We conclude that TFAM phosphorylation by protein kinase A and nonenzymatic acetylation by acetyl-CoA are unlikely to occur at the mitochondrial DNA and that modified free TFAM retains its vital functionalities like compaction and transcription initiation while enhancing transcription processivity.

SUBMITTER: Reardon SD

PROVIDER: S-EPMC9006650 | biostudies-literature | 2022 Apr

REPOSITORIES: biostudies-literature

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Phosphorylation and acetylation of mitochondrial transcription factor A promote transcription processivity without compromising initiation or DNA compaction.

Reardon Sean D SD Mishanina Tatiana V TV

The Journal of biological chemistry 20220310 4

Mitochondrial transcription factor A (TFAM) plays important roles in mitochondrial DNA compaction, transcription initiation, and in the regulation of processes like transcription and replication processivity. It is possible that TFAM is locally regulated within the mitochondrial matrix via such mechanisms as phosphorylation by protein kinase A and nonenzymatic acetylation by acetyl-CoA. Here, we demonstrate that DNA-bound TFAM is less susceptible to these modifications. We confirmed using EMSAs ...[more]

PMID: 35278431

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Project description:Mitochondrial uncoupling proteins (UCPs) are central in the regulation of mitochondrial activity and reactive oxygen species (ROS) production. High oxidative stress is a major cause of male infertility; however, UCPs expression and function in human spermatozoa are still unknown. Herein, we aimed to assess the expression and function of the different homologs (UCP1-6) in human spermatozoa. For this purpose, we screened for the mRNA expression of all UCP homologs. Protein expression and immunolocalization of UCP1, UCP2, and UCP3 were also assessed. Highly motile spermatozoa were isolated from human normozoospermic seminal samples (n = 16) and incubated with genipin, an inhibitor of UCPs (0, 0.5, 5, and 50 µM) for 3 h at 37 °C. Viability and total motility were assessed. Mitochondrial membrane potential and ROS production were evaluated. Media were collected and the metabolic profile and antioxidant potential were analyzed by 1H-NMR and FRAP, respectively. The expression of all UCP homologs (UCP1-6) mRNA by human spermatozoa is herein reported for the first time. UCP1-3 are predominant at the head equatorial segment, whereas UCP1 and UCP2 are also expressed at the spermatozoa midpiece, where mitochondria are located. The inhibition of UCPs by 50 µM genipin, resulting in the UCP3 inhibition, did not compromise sperm cell viability but resulted in irreversible total motility loss that persisted despite washing or incubation with theophylline, a cAMP activator. These effects were associated with decreased mitochondrial membrane potential and lactate production. No differences concerning UCP3 expression, however, were observed in spermatozoa from normozoospermic versus asthenozoospermic men (n = 6). The inhibition of UCPs did not increase ROS production, possibly due to the decreased mitochondrial activity and genipin antioxidant properties. In sum, UCPs are major regulators of human spermatozoa motility and metabolism. The discovery and characterization of UCPs' role in human spermatozoa can shed new light on spermatozoa ROS-related pathways and bioenergetics physiology.

Dataset Information

Phosphorylation and acetylation of mitochondrial transcription factor A promote transcription processivity without compromising initiation or DNA compaction.

Publications

Phosphorylation and acetylation of mitochondrial transcription factor A promote transcription processivity without compromising initiation or DNA compaction.

Similar Datasets

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