Biology subjects

Green, K. M.

Publications and source records attributed to Green, K. M..

4 recordsLinked to original sources

bioRxiv2021

DENR/MCTS1 knockdown modulates repeat-associated non-AUG translation

Repeat associated non-AUG (RAN) translation of mRNAs containing repeat-expansion mutations produces toxic peptides in neurons of patients suffering from neurodegenerative diseases. Recent findings indicate that RAN translation in diverse model systems is not inhibited by cellular stressors that impair global translation through phosphorylation of the alpha subunit of eIF2, the essential eukaryotic translation initiation factor that brings the initiator tRNA to the 40S ribosome. Using in vitro, cell-based, and Drosophila models, we examined the role of alternative ternary complex factors that may function in place of eIF2, including eIF2A, eIF2D, and DENR/MCTS1. Among these factors, DENR knockdown had the greatest inhibitory effect on RAN translation of expanded GGGGCC and CGG repeat reporters, and its reduction improved survival of Drosophila expressing expanded GGGGCC repeats. Taken together, these data support a role for alternative initiation factors in RAN translation and suggest they may serve as novel therapeutic targets in neurodegenerative disease.

neuroscience↗

bioRxiv2021

The RNA helicase DHX36/G4R1 modulates C9orf72 GGGGCC repeat-associated translation

GGGGCC (G4C2) hexanucleotide repeat expansions (HRE) in C9orf72 are the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Repeat-associated non-AUG (RAN) translation of this expansion generates toxic proteins that accumulate in patient brains and contribute to disease pathogenesis. The DEAH-Box Helicase 36 (DHX36/G4R1) plays active roles in RNA and DNA G-quadruplex (G4) resolution in cells. As G4C2 repeats form G4 structures in vitro, we sought to determine the impact of manipulating DHX36 expression on repeat transcription and RAN translation. We found that DHX36 depletion suppresses RAN translation from reporter constructs in a repeat length dependent manner while overexpression of DHX36 enhances RAN translation from G4C2 reporter RNAs. Taken together, these results suggest that DHX36 is active in regulating G4C2 repeat translation, providing potential implications for therapeutic development in nucleotide repeats expansion disorders.

molecular biology↗

bioRxiv2021

In vivo CGG repeat RNA binding protein capture identifies RAN translation modifiers and suppressors of repeat toxicity.

Fragile X-associated tremor/ataxia syndrome (FXTAS) is a neurodegenerative disorder caused by a transcribed CGG repeat expansion in the 5 UTR of FMR1. Expanded CGG repeat RNAs both sequester RNA-binding proteins (RBPs) into nuclear foci and undergo repeat-associated non-AUG (RAN) translation into toxic homopolymeric peptides. RBPs that interact with CGG repeats may play a pivotal role in foci formation and/or RAN translation. Here we employed a CGG repeat RNA-tagging system to capture and identify CGG repeat binding RBPs in vivo under different cellular conditions. We found that several SR (serine/arginine-rich domain) proteins interact with CGG repeat RNAs basally and under cellular stress. These same proteins strongly modify toxicity in a Drosophila model of FXTAS, improving eye degeneration and survival. Furthermore, genetic or pharmacological targeting of the serine/arginine protein kinases (SRPKs) suppresses RAN translation in cellular reporters and toxicity in fly models of FXTAS and C9orf72 ALS/FTD. Finally, pharmacological targeting of SRPK1 supressed CGG repeat toxicity and enhanced survival in rodent neurons. Taken together, these findings demonstrate roles for CGG repeat RNA binding proteins in both RAN translation and repeat toxicity and suggest SRPK inhibition may serve as a possible therapeutic strategy in repeat expansion disorders.

neuroscience↗

bioRxiv2019

Quantifying subclinical and longitudinal microvascular changes following episcleral plaque brachytherapy (EPB) using spectral-domain OCT angiography.

BackgroundI-125 episcleral plaque brachytherapy (EPB) is standard-of-care for globe-conserving treatment of medium-sized choroidal melanomas. Radiation retinopathy is a potential consequence of treatment, characterized by deleterious effects on retinal microvasculature. We investigated the application of Optical Coherence Tomography Angiography (OCTA) for detecting and longitudinally monitoring I-125 episcleral plaque brachytherapy induced radiation retinopathy. MethodsHigh resolution OCTA of the central 3x3mm macula were obtained from I-25 episcleral plaque brachytherapy treated and untreated fellow eyes of 62 patients. Capillary density (vessel skeleton density, VSD) and caliber (vessel diameter index, VDI) were quantified using previously validated semi-automated algorithms. Nonperfusion was also quantified as flow impairment regions (FIR). Exams from treated and fellow eyes obtained pre-treatment and at 6-month, 1-year, and 2-year intervals were compared using generalized estimating equation linear models. Dosimetry maps were used to evaluate spatial correlation between radiation dose and microvascular metrics. ResultsMean time from treatment to last follow-up was 10.8 months. Mean{+/-}SD and median radiation dose at the fovea were 64.5 {+/-} 76 Gy and 32.0 Gy, respectively. Preoperative logMAR (Snellen) mean visual acuity was 0.26 {+/-} 0.05 ([~]20/35) and 0.08 {+/-} 0.02 ([~]20/25) in treated and fellow eyes, respectively. At 6 months, treated eyes had significantly lower VSD (0.147 {+/-} 0.003 vs 0.155 {+/-} 0.002; p = 0.023) and higher FIR (1.95 {+/-} 0.176 vs 1.45 {+/-} 0.099; p = 0.018) compared to fellow eyes. There was a significant decrease in VSD and a corresponding increase in FIR even for treated eyes without clinically identifiable retinopathy at 6 months. VDI was significantly higher in treated eyes than in fellow eyes at 2 years (2.93 {+/-} 0.022 vs 2.84 {+/-} 0.016; p = 0.002). Microvascular changes were spatially correlated with a radiation gradient of 85-250 Gy across the fovea. ConclusionsOCTA can be used to quantify and monitor EPB induced radiation, and can detect vascular abnormalities even in the absence of clinically observable retinopathy. OCTA may therefore be useful in investigating treatment interventions that aim to delay EPB-induced radiation retinopathy.

scientific communication and education↗