a-colorimetric-assay-citrate-synthase-activity-drosophila

A Colorimetric Assay of Citrate Synthase Activity in <em>Drosophila Melanogaster</em>

Mitochondria play the most prominent roles in cellular metabolism by producing ATP through oxidative phosphorylation and regulating a variety of ...

crispr-based-modular-assembly-for-high-throughput-construction-uas

CRISPR-Based Modular Assembly for High-Throughput Construction of a UAS-cDNA/ORF Plasmid Library

Functional genomics screening offers a powerful approach to probe gene function and relies on the construction of genome-wide plasmid libraries. ...

embryo-microinjection-for-transgenesis-in-drosophila

Embryo Microinjection for Transgenesis in <em>Drosophila</em>

Transgenesis in Drosophila is an essential approach to studying gene function at the organism level. Embryo microinjection is a crucial step for the ...

crispr-based-shuttle-cloning-a-high-throughput-cloning-method

CRISPR-based Shuttle Cloning: A High-throughput Cloning Method

The development of genome-wide plasmid libraries using existing genomic repositories serves as a pivotal prerequisite for systematic functional ...

The transcriptional co-activator PGC-1α is a key regulator of mitochondrial function and muscle fiber specification in the skeletal muscle. The E3 ubiquitin ligase RNF34 ubiquitinates PGC-1α and negatively regulates mammalian brown fat cell metabolism. However, the functional importance of RNF34 in the skeletal muscle and its impact on energy metabolism remain unknown. The Drosophila PGC-1 homolog dPGC-1 and its mammalian counterparts have conserved functions in mitochondria and insulin signaling. Here, we showed that the Drosophila RNF34 (dRNF34) ubiquitinates the Drosophila PGC-1α (dPGC-1) and promotes its degradation in HEK293T cells by immunoprecipitation and western blot analysis. This allows us to use Drosophila as a powerful model system to study the physiological role of RNF34 in mitochondrial function and metabolism. In the in vivo studies, by separately expressing two independent UAS-dRNF34 RNAi transgenes driven by the muscle-specific 24B-Gal4 driver, we found that knockdown of dRNF34 specifically in muscle promotes mitochondrial biogenesis, improves negative geotaxis, extends climbing time to exhaustion in moderate aged flies and counteracts high-fat-diet-induced high triglyceride content. Furthermore, we showed that knockdown of dPGC-1 reversed the effects of the dRNF34 knockdown phenotypes described above. Our results reveal that dRNF34 plays an important role in regulating mitochondrial biogenesis in muscle and lipid metabolism through dPGC-1. Thus, inhibition of RNF34 activity provides a potential novel therapeutic strategy for the treatment of age-related muscle dysfunction.

RNF34 modulates the mitochondrial biogenesis and exercise capacity in muscle and lipid metabolism through ubiquitination of PGC-1 in Drosophila.

Pathological TDP-43 aggregation is characteristic of several neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD-TDP); however, how TDP-43 aggregation and function are regulated remain poorly understood. Here, we show that O-GlcNAc transferase OGT-mediated O-GlcNAcylation of TDP-43 suppresses ALS-associated proteinopathies and promotes TDP-43's splicing function. Biochemical and cell-based assays indicate that OGT's catalytic activity suppresses TDP-43 aggregation and hyperphosphorylation, whereas abolishment of TDP-43 O-GlcNAcylation impairs its RNA splicing activity. We further show that TDP-43 mutations in the O-GlcNAcylation sites improve locomotion defects of larvae and adult flies and extend adult life spans, following TDP-43 overexpression in Drosophila motor neurons. We finally demonstrate that O-GlcNAcylation of TDP-43 promotes proper splicing of many mRNAs, including STMN2, which is required for normal axonal outgrowth and regeneration. Our findings suggest that O-GlcNAcylation might be a target for the treatment of TDP-43-linked pathogenesis.

ArticleTotal : 4	Year
Publication title Cited by 1	2020
Publication title Cited by 1	2024
Publication title	2024
Publication title	2025

Article	Year
RNF34 modulates the mitochondrial biogenesis and exercise capacity in muscle and lipid metabolism through ubiquitination of PGC-1 in Drosophila. Acta biochimica et biophysica Sinica\| PubMed ID: 30247505	2018
O-GlcNAcylation of TDP-43 suppresses proteinopathies and promotes TDP-43's mRNA splicing activity. EMBO reports\| PubMed ID: 33855783	2021
CRISPR-based shuttle cloning of 1,397 human genes into UAS vectors. Acta biochimica et biophysica Sinica\| PubMed ID: 40285452	2025

Jiwu Wang

Biography

JoVE Journal Publications

Other Publications