ABSTRACT
A hallmark of aging is loss of differentiated cell identity. Aged Drosophila midgut differentiated enterocytes (ECs) lose their identity, impairing tissue homeostasis. To discover identity regulators, we performed an RNAi screen targeting ubiquitin-related genes in ECs. Seventeen genes were identified, including the deubiquitinase Non-stop (CG4166). Lineage tracing established that acute loss of Non-stop in young ECs phenocopies aged ECs at cellular and tissue levels. Proteomic analysis unveiled that Non-stop maintains identity as part of a Non-stop identity complex (NIC) containing E(y)2, Sgf11, Cp190, (Mod) mdg4, and Nup98. Non-stop ensured chromatin accessibility, maintaining the EC-gene signature, and protected NIC subunit stability. Upon aging, the levels of Non-stop and NIC subunits declined, distorting the unique organization of the EC nucleus. Maintaining youthful levels of Non-stop in wildtype aged ECs safeguards NIC subunits, nuclear organization, and suppressed aging phenotypes. Thus, Non-stop and NIC, supervise EC identity and protects from premature aging.
Subject(s)
Aging, Premature/genetics , Aging/genetics , Drosophila Proteins/genetics , Drosophila melanogaster/physiology , Enterocytes/physiology , Animals , Disease Models, Animal , Drosophila Proteins/metabolism , Female , Male , Phenotype , ProteomeABSTRACT
The Spt-Ada-Gcn5 Acetyltransferase (SAGA) chromatin modifying complex is a critical regulator of gene expression and is highly conserved across species. Subunits of SAGA arrange into discrete modules with lysine aceyltransferase and deubiquitinase activities housed separately. Mutation of the SAGA deubiquitinase module can lead to substantial biological misfunction and diseases such as cancer, neurodegeneration, and blindness. Here, we review the structure and functions of the SAGA deubiquitinase module and regulatory mechanisms acting to control these.
Subject(s)
Deubiquitinating Enzymes/metabolism , Multienzyme Complexes/metabolism , Trans-Activators/metabolism , Transcriptional Activation , p300-CBP Transcription Factors/metabolism , Animals , Arabidopsis/enzymology , Aspergillus nidulans/enzymology , Ataxin-7/genetics , Blindness/genetics , Deubiquitinating Enzymes/genetics , Drosophila/enzymology , Histones/metabolism , Humans , Mice , Multienzyme Complexes/genetics , Mutation , Neoplasms/genetics , Neurodegenerative Diseases/genetics , Peptides/genetics , Protein Processing, Post-Translational , RNA Polymerase II/metabolism , Saccharomyces cerevisiae/enzymology , Trans-Activators/genetics , p300-CBP Transcription Factors/geneticsABSTRACT
Nearly universal among organisms, circadian rhythms coordinate biological activity to earth's orbit around the sun. To identify factors creating this rhythm and to understand the resulting outputs, entrainment of model organisms to defined circadian time-points is required. Here we detail a procedure to entrain many Drosophila to a defined circadian rhythm. Furthermore, we detail post-entrainment steps to prepare samples for immunofluorescence, nucleic acid, or protein extraction-based analysis.
