Biology subjects

Holder, J.

Publications and source records attributed to Holder, J..

6 recordsLinked to original sources

bioRxiv2026

Selective miniprotein inhibitors of Aurora-A kinase designed using interaction-motif scaffolding

Targeting kinase ATP-binding sites has produced many successful therapeutics, but selectivity remains a major challenge. We hypothesised that substrate-recognition surfaces could provide an alternative route to selective kinase inhibition. Here, we designed and tested structure-guided miniprotein inhibitors of Aurora-A using N-Myc as a template, a natural weak binder of the P+1 pocket. The most potent designs bind Aurora-A with single-digit nanomolar affinity, more than a thousand fold higher than the starting template, and selectively inhibit Aurora-A over Aurora-B in kinase assays and mitotic cells. Despite diverse architectures, successful binders targeting the substrate-recognition surface converged on a common strategy, coupling extensive engagement of the G helix with activation loop stabilisation. Together these results show that selective kinase inhibition can be achieved by exploiting structurally well-defined active-state conformations rather than kinase-specific inactive states. Our work establishes a framework for converting weak substrate-recognition surface interactors into potent, selective kinase inhibitors through structure-guided protein design.

molecular biology↗

bioRxiv2024

A novel splicing graph allows a direct comparison between exon-based and splice junction-based approaches to alternative splicing detection

There are primarily two computational approaches to alternative splicing detection: splice junction-based and exon-based approaches. Despite their shared goal of addressing the same biological problem, these approaches have not been reconciled before. We devised a novel graph structure and algorithm aimed at mapping between the exonic parts and splicing events detected by the two different methods. Through simulations, we demonstrated disparities in sensitivity and specificity between splice junction-based and exon-based methods. When applied to empirical data, there were large discrepancies in the results, suggesting that the methods are complementary. With the discrepancies localized to individual events and exonic parts, we were able to gain insights into the strengths and weaknesses inherent in each approach. Finally, we integrated the results to generate a comprehensive list of both common and unique alternative splicing events detected by both methodologies. Availabilityhttps://github.com/HanLabUNLV/GrASE Contactmira.han@unlv.edu Supplementary informationSupplementary data are available online.

bioinformatics↗

bioRxiv2022

Speaker-induced suppression in EEG during a naturalistic reading and listening task

Speaking elicits a suppressed neural response when compared to listening to others speech, a phenomenon known as speaker-induced suppression (SIS). Previous research has focused on investigating SIS at constrained levels of linguistic representation, such as the individual phoneme and word level. Here we present scalp EEG data from a dual speech perception and production task where participants read sentences aloud then listened to playback of themselves reading those sentences. Playback was separated into predictable repetition of the previous trial and unpredictable, randomized repetition of a former trial to investigate the role predictive processing plays in SIS. Concurrent EMG was recorded to control for movement artifact during speech production. In line with previous research, event-related potential analyses at the sentence level demonstrated suppression of early auditory components of the EEG for production compared to perception. To evaluate whether specific neural representations contribute to SIS (in contrast with a global gain change), we fit linear encoding models that predicted scalp EEG based on phonological features, EMG activity, and task condition. We found that phonological features were encoded similarly between production and perception. However, this similarity was only observed when controlling for movement by using the EMG response as an additional regressor. Our results suggest SIS is at the representational level a global gain change between perception and production, not the suppression of specific characteristics of the neural response. We also detail some important considerations when analyzing EEG during continuous speech production.

neuroscience↗

bioRxiv2021

Generalizable EEG encoding models with naturalistic audiovisual stimuli

In natural conversations, listeners must attend to what others are saying while ignoring extraneous background sounds. Recent studies have used encoding models to predict electroencephalography (EEG) responses to speech in noise-free listening situations, sometimes referred to as "speech tracking" in EEG. Researchers have analyzed how speech tracking changes with different types of background noise. It is unclear, however, whether neural responses from noisy and naturalistic environments can be generalized to more controlled stimuli. If encoding models for noisy, naturalistic stimuli are generalizable to other tasks, this could aid in data collection from populations who may not tolerate listening to more controlled, less-engaging stimuli for long periods of time. We recorded non-invasive scalp EEG while participants listened to speech without noise and audiovisual speech stimuli containing overlapping speakers and background sounds. We fit multivariate temporal receptive field (mTRF) encoding models to predict EEG responses to pitch, the acoustic envelope, phonological features, and visual cues in both noise-free and noisy stimulus conditions. Our results suggested that neural responses to naturalistic stimuli were generalizable to more controlled data sets. EEG responses to speech in isolation were predicted accurately using phonological features alone, while responses to noisy speech were more accurate when including both phonological and acoustic features. These findings may inform basic science research on speech-in-noise processing. Ultimately, they may also provide insight into auditory processing in people who are hard of hearing, who use a combination of audio and visual cues to understand speech in the presence of noise. Significance StatementUnderstanding spoken language in natural environments requires listeners to parse acoustic and linguistic information in the presence of other distracting stimuli. However, most studies of auditory processing rely on highly controlled stimuli with no background noise, or with background noise inserted at specific times. Here, we compare models where EEG data are predicted based on a combination of acoustic, phonetic, and visual features in highly disparate stimuli - sentences from a speech corpus, and speech embedded within movie trailers. We show that modeling neural responses to highly noisy, audiovisual movies can uncover tuning for acoustic and phonetic information that generalizes to simpler stimuli typically used in sensory neuroscience experiments.

neuroscience↗

bioRxiv2020

PP1 promotes cyclin B destruction and the metaphase-anaphase transition by dephosphorylating CDC20

Ubiquitin-dependent proteolysis of cyclin B and securin initiates sister chromatid segregation and anaphase. The anaphase promoting complex/cyclosome (APC/C) and its co-activator CDC20 form the main ubiquitin E3 ligase for these proteins. APC/CCDC20 is regulated by CDK1-cyclin B and counteracting PP1 and PP2A family phosphatases through modulation of both activating and inhibitory phosphorylations. Here we report that PP1 promotes cyclin B destruction at the onset of anaphase by removing specific inhibitory phosphorylation in the N-terminus of CDC20. Depletion or chemical inhibition of PP1 stabilises cyclin B and results in a pronounced delay at the metaphase-to-anaphase transition after chromosome alignment. This requirement for PP1 is lost in cells expressing CDK1-phosphorylation defective CDC206A mutants. These CDC206A cells show a normal spindle checkpoint response, but once all chromosomes have aligned rapidly degrade cyclin B and enter into anaphase in the absence of PP1 activity. PP1 therefore facilitates the metaphase-to-anaphase by promoting APC/CCDC20-dependent destruction of cyclin B in human cells.

cell biology↗

bioRxiv2020

Ordered dephosphorylation initiated by the selective proteolysis of cyclin B drives mitotic exit

APC/C-mediated proteolysis of cyclin B and securin promotes entry into anaphase, inactivating CDK1 and permitting chromosome segregation, respectively. Reduction of CDK1 activity relieves inhibition of the CDK1-opposing phosphatases PP1 and PP2A-B55 leading to dephosphorylation of substrates crucial for mitotic exit. Meanwhile, continued APC/C activity is required to target various proteins, including Aurora and Polo kinases, for degradation. Together, these activities orchestrate a complex series of events during mitotic exit. However, the relative importance of regulated proteolysis and dephosphorylation in dictating the order and timing of these events remains unclear. Using high temporal-resolution mass spectrometry, we compare the relative extent of proteolysis and protein dephosphorylation. This reveals highly-selective rapid ([~]5min half-life) proteolysis of cyclin B, securin and geminin at the metaphase to anaphase transition, followed by slow proteolysis (>60 min half-life) of other mitotic regulators. Protein dephosphorylation requires APC/C-dependent destruction of cyclin B and was resolved into PP1-dependent fast, intermediate and slow categories with unique sequence motifs. We conclude that dephosphorylation initiated by the selective proteolysis of cyclin B drives the bulk of changes observed during mitotic exit.

cell biology↗