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Principal Investigator: ANITA H. CORBETT
Organization: EMORY UNIVERSITY
Fiscal Year: 2024
Award: $369,822
Funding agency: National Institute of Neurological Disorders and Stroke
Project Summary
Post-transcriptional control of mRNAs plays a key role in regulating important developmental events via RNA
binding proteins (RBPs) that modulate key aspects of RNA fate, including splicing, stability, and translation.
Developmental roles of RBPs are particularly evident in the nervous system, where their loss is linked to an array
of diseases. Understanding how individual RBPs contribute to tissue-specific developmental events is
complicated by their generally ubiquitous expression and by the insufficiency of consensus RBP-binding sites to
predict in vivo occupancy or regulation. These observations imply as yet undefined mechanisms that elevate
requirements for RBPs in specific cell types. We co-discovered an inherited neurodevelopmental disease caused
by loss of the ubiquitously expressed, zinc finger RBP ZC3H14 and have probed function of this protein in
multiple model organisms. The ZC3H14 ortholog in D. melanogaster, Nab2 (nuclear polyadenylated RNA binding
protein 2), provides a tractable genetic model to probe molecular and neurodevelopmental roles. Importantly,
ZC3H14/Nab2 is expressed in all cells but required specifically in neurons of the developing nervous system to
support viability, brain axon guidance, locomotion, and olfactory memory. Critically, human ZC3H14 can
substitute for Nab2 in fly neurons, indicating that the two orthologs share molecular roles and target RNAs.
However, the identity of ZC3H14/Nab2-regulated RNAs, and mechanisms that elevate the ZC3H14/Nab2 role in
neurons, are gaps that limit understanding of how such RBPs play key roles in neurons. Our recent work
exploiting Drosophila provides a foundation to fill these gaps by identifying Nab2 as a novel inhibitor of N6-
methyladenosine (m6A) on a subset of mRNAs in the developing nervous system. m6A is deposited at specific
sites within mRNAs by the m6A ‘writer’ Mettl3. m6A is enriched in the developing brain, where it plays key roles
in neurodevelopment. Our analysis of one newly identified Nab2 neuronal target, the pre-mRNA encoding the
Sex Lethal (Sxl) sex determination factor, suggests that Nab2 promotes a key developmentally regulated exon-
skipping event in Sxl pre-mRNA by binding an A-rich intronic element and inhibiting m6A deposition by Mettl3.
Critically, Nab2-regulation of this splicing event is neuron-specific and rescue of multiple Nab2 phenotypes by
concurrent Mettl3 loss implies that the Nab2-m6A hypermethylation model extends to additional RNA targets.
We hypothesize that the neurodevelopmental requirement for Nab2/ZC3H14 is based on regulating on a set
of mRNAs encoding key neuronal proteins, at least in part, via this novel m6A inhibitory role. We pursue this
mechanistic hypothesis in three aims: Aim 1) Exploit the Sxl pre-mRNA as a model to dissect Nab2 inhibition of
m6A deposition; Aim 2) Use a Sxl splicing reporter to parse direct and direct effects of Nab2 and m6A-regulation
of splicing and identify additional factors in the Nab2-m6A pathway; and Aim 3) Identify key neurodevelopmental
targets of Nab2/ZC3H14 in both flies and in vertebrates. Taken together, these approaches will provide insight
into why loss of a ubiquitously expressed RNA binding protein causes morphogenesis defects in the brain.
Terms: <AICDA><AICDA protein><AID gene><AID protein><Affect><Assay><Behavioral><Binding><Binding Proteins><Binding Sites><Bioassay><Biologic Models><Biological Assay><Biological Models><Body Tissues><Brain><Brain Nervous System><CDA2 protein><CNS Nervous System><Cell Body><Cell Line><CellLine><Cells><Central Nervous System><Cis-Acting Locus><Cis-Acting Sequence><Clinical><Combining Site><Consensus><Coupled><Data><Defect><Dendritic Spines><Deposit><Deposition><Development><Disease><Disorder><Drosophila><Drosophila genus><Drosophila melanogaster><Elements><Encephalon><Event><Exons><Experimental Genetics><Female><Flies><Foundations><GeneHomolog><Genes><Genetic><Genetic Alteration><Genetic Change><Genetic Models><Genetic defect><Hereditary><Heritability><Homolog><Homologous Gene><Homologue><Human><Hypermethylation><In vivo analysis><Individual><Inherited><Intellectual disability><Intellectual functioning disability><Intellectual limitation><Intervening Sequences><Introns><Ligand Binding Protein><Ligand Binding Protein Gene><Link><Locomotion><Maps><Mediating><Memory><Messenger RNA><Mice><Mice Mammals><Model System><Modeling><Modern Man><Modification><Molecular><Molecular Interaction><Molecular Probes><Morphogenesis><Morphology><Murine><Mus><Mutation><Nerve Cells><Nerve Unit><Nervous System><Neural Cell><Neural Development><Neuraxis><Neurocyte><Neurodevelopmental Disorder><Neurologic Body System><Neurologic Organ System><Neurological Development Disorder><Neuronal Differentiation><Neurons><Non-Polyadenylated RNA><Nuclear><Organ><Ortholog><Orthologous Gene><Pathway interactions><Phenotype><Play><Poly(A)+ RNA><Polyadenylated RNA><Post-Transcriptional Control><Post-Transcriptional Regulation><Pre-mRNA><Protein Binding><Proteins><Public Health><Publishing><RNA><RNA Gene Products><RNA Splicing><RNA, Messenger, Precursors><RNA-Binding Proteins><Reactive Site><Reader><Regulation><Reporter><Ribonucleic Acid><Role><Site><Specific qualifier value><Specified><Splicing><Strains Cell Lines><Testing><Tissues><Trans-Acting Factors><Trans-Activators><Transactivators><Transcript><Translations><Vertebrate Animals><Vertebrates><Work><Zinc Finger Domain><Zinc Finger Motifs><Zinc Fingers><activation-induced cytidine deaminase><activation-induced deaminase><axon growth cone guidance><axon guidance><bound protein><cell type><cultured cell line><dendrite spine><developmental><exon skipping><fly><fruit fly><genome mutation><global gene expression><global transcription profile><human disease><in vivo><in vivo evaluation><in vivo testing><inhibitor><insight><intellectual and developmental disability><limited intellectual functioning><mRNA><mRNA Precursor><model organism><morphogenetic process><neurodevelopment><neurodevelopmental disease><neuronal><novel><pathway><polyadenosine><polyriboadenosine><post-transcriptional gene regulation><posttranscriptional control><posttranscriptional regulation><protein function><recruit><sex><sex determination><social role><tool><transcriptome><translation><unpublished works><vertebrata>