Biology subjects

Greenblatt, E. J.

Publications and source records attributed to Greenblatt, E. J..

4 recordsLinked to original sources

bioRxiv2026

FMRP prevents Me31B/DDX6-associated repression of long neurodevelopmental mRNAs

Long, dosage-sensitive mRNAs encoding neurodevelopmental regulators are selectively dependent on fragile X messenger ribonucleoprotein (FMRP) for efficient translation across animal systems, but the mechanism underlying this length-dependent requirement remains unclear. Here, we show that FMRP maintains translation of long target mRNAs by preventing their inappropriate sequestration into a Me31B/DDX6-dependent P-body repression pathway. In Drosophila oocytes, loss of FMRP caused target mRNAs, but not bulk polyadenylated RNA, to accumulate in Me31B-marked P-bodies. Improved individual-nucleotide resolution crosslinking and immunoprecipitation (iiCLIP) revealed that FMRP and Me31B co-occupy many long coding sequences, suggesting that FMRP targets are intrinsically vulnerable to repressive Me31B-associated machinery. A screen targeting [~]120 candidate genes, followed by proteomic analysis, identified an FMRP-associated ribonucleoprotein (RNP) assembly containing the stress granule-linked proteins Rin/G3BP, Lig/UBAP2L, and Capr/Caprin1. These factors supported translation or localization of distinct FMRP target subsets. Finally, inhibition of Me31B-dependent repressive complex assembly restored translation of the majority of FMRP targets in FMRP-deficient oocytes, supporting a model in which translational failure results from excessive repression. Together, these findings reveal a cytoplasmic RNP assembly that safeguards mRNAs by antagonizing promiscuous P-body repression and provide a mechanistic explanation for the selective vulnerability of long neurodevelopmental transcripts to FMRP loss.

molecular biology↗

bioRxiv2020

Fmr1 translationally activates stress-sensitive mRNAs encoding large proteins in oocytes and neurons

Mutations in Fmr1, encoding the RNA binding protein FMRP, are leading causes of intellectual disability, autism, and female infertility, but FMRPs mechanism of action is controversial. In contrast to its previously postulated function as a translation repressor acting by stalling elongation, we recently found that FMRP activates translation initiation of large proteins in Drosophila oocytes up to [~]2-fold. We report here that FMRPs function as a translational activator is conserved in the mammalian brain. Reanalysis of mouse cortex ribosome profiling data shows that translation of large proteins in Fmr1 mutants is down-regulated 2.0-1.2-fold; ribosome stalling appears not to influence FMRP target protein translation in either cortex or oocyte tissue. Consistent with an activator function, most FMRP targets are associated with clinical syndromes when reduced, but not when over-expressed. Fmr1-dependent translation of one target, the N-end rule E3 ligase Poe/UBR4, occurs in microscopically visible ribonucleoprotein particles. These "Poe particles" require FMRP for their formation, are distinct from P bodies, and depend on actively elongating ribosomes, as indicated by their dissolution following a brief puromycin treatment. N-end rule-mediated proteolysis via Poe/UBR4 restrains cell growth and limits MAPK signaling in nervous tissue. Thus, loss of FMRP reduces production of an important growth repressor.

neuroscience↗

bioRxiv2019

Prolonged ovarian storage of mature Drosophila oocytes dramatically increases meiotic spindle instability

More than 95% of fertilized Drosophila oocytes from outbred stocks develop fully regardless of maternal age, in contrast to human oocytes, which frequently generate non-viable aneuploid embryos. Since Drosophila oocytes are normally stored only briefly prior to ovulation, unlike their human counterparts, we investigated the effects of storage on oocyte quality. Using a novel system to acquire oocytes held for known periods, we analyzed by ribosome profiling how translation and cellular function change over time. Oocyte developmental capacity decays in a precise temperature-dependent manner over 1-4 weeks, due to a progressive inability to complete meiosis. Meiotic metaphase genes, the Fmr1 translational regulator, and the small heat shock protein chaperones Hsp26 and Hsp27 are preferentially translated during storage, and oocytes lacking Hsp26 and Hsp27 age prematurely. However translation falls generally 2.3-fold with age despite constant mRNA levels, and this inability to maintain translational equilibrium correlates with oocyte functional decline. These findings show that meiotic chromosome segregation in Drosophila oocytes is uniquely sensitive to prolonged quiescence, and suggest that the extended storage of mature human oocytes contributes to their chromosome instability. If so, then these problems may be more amenable to intervention than previously supposed.

developmental biology↗

bioRxiv2018

Fragile X mental retardation protein is a size-dependent translational activator

FMR1 enhances translation of large neural/oocyte proteins\n\nMutations in the highly conserved Fragile X mental retardation gene (Fmr1) cause the most common inherited human intellectual disability/autism spectrum disorder. Fmr1 is also needed for ovarian follicle development, and lesions are the largest genetic cause of premature ovarian failure (POF). FMR1 associates with ribosomes and is thought to repress translation, but identifying functional targets has been difficult. We analyzed FMR1s role in quiescent Drosophila oocytes stored prior to ovulation, cells that depend entirely on translation of stored mRNA. Ribosome profiling revealed that in quiescent oocytes FMR1 stimulates the translation of large proteins, including at least twelve proteins whose human homologs are associated with dominant intellectual disability disorders, and 25 others associated with neural dysfunction. Knockdown of Fmr1 in unstored oocytes did not affect embryo development, but more than 50% of embryos derived from stored oocytes lacking FMR1 developed severe neural defects. Fmr1s previously unappreciated role promoting the translation of large proteins from stored mRNAs in oocytes and neurons may underlie POF as well as multiple aspects of neural dysfunction.

developmental biology↗