The Arf-GAPs, AoAge1 and AoAge2, regulate diverse cellular processes, conidiation, trap formation, and secondary metabolism in Arthrobotrys oligospora.

Guanine nucleotide-binding proteins of the ADP ribosylation factor (Arf) family and their activating proteins (Arf-GAPs) are essential for diverse biological processes. Here, two homologous Arf-GAPs, Age1 (AoAge1) and Age2 (AoAge2), were identified in the widespread nematode-trapping fungus Arthrobotrys oligospora. Our results demonstrated that AoAge1, especially AoAge2, played crucial roles in mycelial growth, sporulation, trap production, stress response, mitochondrial activity, DNA damage, endocytosis, reactive oxygen species production, and autophagy. Notably, transcriptome data revealed that approximately 62.7% of the genes were directly or indirectly regulated by AoAge2, and dysregulated genes in Aoage2 deletion were enriched in metabolism, ribosome biogenesis, secondary metabolite biosynthesis, and autophagy. Furthermore, Aoage2 inactivation caused a substantial reduction in several compounds compared to the wild-type strain. Based on these results, a regulatory network for AoAge1 and AoAge2 was proposed and verified using a yeast two-hybrid assay. Based on our findings, AoAge1 and AoAge2 are essential for vegetative growth and mycelial development. Specifically, AoAge2 is required for sporulation and trapping morphogenesis. Our results demonstrated the critical functions of AoAge1 and AoAge2 in mycelial growth, diverse cellular processes, and pathogenicity, offering deep insights into the functions and regulatory mechanisms of Arf-GAPs in nematode-trapping fungi.

Length Polymorphism and Methylation Status of UPS29 Minisatellite of the ACAP3 Gene as Molecular Biomarker of Epilepsy. Sex Differences in Seizure Types and Symptoms.

Epilepsy is a neurological disease with different clinical forms and inter-individuals heterogeneity, which may be associated with genetic and/or epigenetic polymorphisms of tandem-repeated noncoding DNA. These polymorphisms may serve as predictive biomarkers of various forms of epilepsy. ACAP3 is the protein regulating morphogenesis of neurons and neuronal migration and is an integral component of important signaling pathways. This study aimed to carry out an association analysis of the length polymorphism and DNA methylation of the UPS29 minisatellite of the ACAP3 gene in patients with epilepsy. We revealed an association of short UPS29 alleles with increased risk of development of symptomatic and cryptogenic epilepsy in women, and also with cerebrovascular pathologies, structural changes in the brain, neurological status, and the clinical pattern of seizures in both women and men. The increase of frequency of hypomethylated UPS29 alleles in men with symptomatic epilepsy, and in women with both symptomatic and cryptogenic epilepsy was observed. For patients with hypomethylated UPS29 alleles, we also observed structural changes in the brain, neurological status, and the clinical pattern of seizures. These associations had sex-specific nature similar to a genetic association. In contrast with length polymorphism epigenetic changes affected predominantly the long UPS29 allele. We suppose that genetic and epigenetic alterations UPS29 can modify ACAP3 expression and thereby affect the development and clinical course of epilepsy.

Key roles of Arf small G proteins and biosynthetic trafficking for animal development.

Although biosynthetic trafficking can function constitutively, it also functions specifically for certain developmental processes. These processes require either a large increase to biosynthesis or the biosynthesis and targeted trafficking of specific players. We review the conserved molecular mechanisms that direct biosynthetic trafficking, and discuss how their genetic disruption affects animal development. Specifically, we consider Arf small G proteins, such as Arf1 and Sar1, and their coat effectors, COPI and COPII, and how these proteins promote biosynthetic trafficking for cleavage of the Drosophila embryo, the growth of neuronal dendrites and synapses, extracellular matrix secretion for bone development, lumen development in epithelial tubes, notochord and neural tube development, and ciliogenesis. Specific need for the biosynthetic trafficking system is also evident from conserved CrebA/Creb3-like transcription factors increasing the expression of secretory machinery during several of these developmental processes. Moreover, dysfunctional trafficking leads to a range of developmental syndromes.

Simple in vitro assay of Arf GAPs and preparation of Arf proteins as substrates.

Defining the interaction of Arf GAPs with specific Arfs is important for understanding their functions in the endocytic system. Cell-based approaches have been valuable for identifying Arfs and Arf GAPs active in the endocytic compartment; however, the cell-based assays have some limitations in establishing relationships among the Arfs and ArfGAPs. Here we describe a simple in vitro assay that will provide a means for comparing Arfs as substrates and serve to complement cell-based studies.