Biology subjects

Miller, C. A.

Publications and source records attributed to Miller, C. A..

At least 19 recordsLinked to original sources

bioRxiv2026

Long read sequencing reveals novel isoforms and spliceosome-mutant-enriched transcripts in AML and MDS

The alternative splicing landscape of the leukemia transcriptome remains poorly characterized, since short-read sequencing cannot resolve complete transcript structures. Using the Oxford Nanopore cDNA platform, we generated nearly 2 billion long reads (median 25.8 million per sample) from 71 human samples, including 48 acute myeloid leukemia or myelodysplastic syndrome samples, 25 of which had splicing-factor gene mutations (in SRSF2, U2AF1, or SF3B1). An additional 23 samples were from sorted hematopoietic cell populations from healthy individuals. Using this dataset, we created a transcript assembly containing 174,162 novel isoforms that are not described in the reference transcriptome. Deep-scale proteomic validation confirmed that many of these transcripts are translated into protein. We also identified isoforms enriched in spliceosome-mutant samples and found proteomic evidence of frequent nonsense-mediated decay regulation of novel transcripts. This dataset is a valuable community resource, enabling detection of new transcripts in short-read data sets. An interactive portal to explore splicing patterns in these data is available at https://leylab.org/isoforms/. Key PointsLong-read sequencing enables the detection of many novel transcripts in AML, including many from splice-factor-mutant patient samples This expanded transcriptome is a valuable community resource and can be used to improve analyses of short-read RNAseq data

cancer biology↗

bioRxiv2026

Syngap1 Synchronizes Relative Neuronal Maturation Across Cortical Areas to Organize Distributed Functional Networks

Neurodevelopmental disorders are increasingly viewed as disorders of distributed brain networks, yet how developmental perturbations generate co-occurring hypo- and hyper-functional network states remains unclear. We show that Syngap1 haploinsufficiency in mice produces opposing cortical activity patterns: sensory-evoked responses exhibit reduced gain, whereas state-linked movement/arousal activity is amplified. Restricting Syngap1 deficiency to developing cortical excitatory neurons reproduces distributed sensory hypofunction but not movement-linked hyperfunction, indicating non-uniform, cell-type-dependent contributions to these patterns. During a postnatal window of circuit assembly, Layer 2/3 intratelencephalic populations across cortical regions occupy distinct maturation states, reflected in separable dendritic architectures and intrinsic excitability. Perturbing Syngap1 reduces this developmental separability through region-dependent, opposite-direction effects on dendritic maturation and inversion of ERK-sensitive control of intrinsic excitability. We propose that coordinated relative maturation of interacting neuronal populations establishes distributed network balance, and that gene-dependent shifts in this coordination produce stable, opposing network states.

neuroscience↗

bioRxiv2026

Binding Entropy Can Be Predicted by Crystallographic Ensembles

Protein-ligand binding is governed by free energy, comprising both enthalpic and entropic contributions. Yet structural interpretations of binding thermodynamics have predominantly focused on enthalpic interactions, largely neglecting entropy because it is difficult to quantify from static structural models. Here, we developed multiconformer ensemble models to analyze high-resolution X-ray crystallography structures and estimate both protein and solvent conformational entropies. These ensemble models successfully predicted experimental binding entropies measured by isothermal titration calorimetry for over 70 protein-ligand pairs across 12 proteins, revealing a strong linear correlation. Protein entropy, estimated using crystallographic order parameters that capture both harmonic and anharmonic motion, correlates linearly with experimental binding entropy. Incorporating resolution-corrected differences in water-molecule counts substantially improves predictions, demonstrating that protein and solvent contributions must be considered jointly. Analysis of water-protein hydrogen bonding networks partially explains entropic differences across complexes. These results establish that crystallographic ensembles can quantify binding entropy, enabling explicit entropic considerations in structure-based studies of molecular recognition for both functional analysis and drug design.

biophysics↗

bioRxiv2025

Early overactivation of non-muscle myosin II during adaptation to combined BRAF and MEK inhibitors in dedifferentiating cutaneous melanomas

Cutaneous melanoma is a very aggressive type of skin cancer with remarkable phenotypic plasticity that contributes to adaptation and resistance to targeted therapies against the MAPK pathway. Previous research described that non-muscle myosin II (NMII) of the actomyosin cytoskeleton, which is essential for cell migration and metastasis, is overactivated in BRAF inhibitor-resistant melanomas. Since the combination of BRAF and MEK inhibitors (BMi) is the current standard of care, we investigated if and how NMII activity is regulated during adaptation to BMi. Here, we find that most dedifferentiating BMi-resistant melanomas overactivate NMII compared to their parental counterparts. NMII activity generally increases during the first 2 weeks of BMi treatment, and it is followed by elevated total NMII levels due partly to transcriptional modulation. Although ERK activity rebounds with similar kinetics, NMII overactivation is not prevented by ERK inhibition but by blockade of ROCK. In melanomas that hyperdifferentiate during adaptation to BMi, NMII activity is not increased upon BMi treatment due, in part, to MITF. We also find that co-targeting NMII along BMi in some melanomas reduces survival of drug-tolerant persister cells, which would delay the development of resistance. Therefore, our study identifies elevated NMII activity as a potential marker of adaptation to MAPK in some melanoma subpopulations, and also provide an approach to delay the emergence of resistance to MAPK-targeted therapy.

cancer biology↗

bioRxiv2025

Mesoscale proximity labeling to study macro changes to chromatin occupancy

Proximity labeling traditionally identifies interactomes of a single protein or RNA, though this approach limits mechanistic understanding of biomolecules functioning within complex systems. Here, we demonstrate a strategy for deciphering ligand-induced changes to global biomolecular interactions by enabling proximity labelling at the mesoscale, across an entire cellular system. By inserting nanoscale proximity labelling catalysts throughout chromatin, this system, MesoMap, provided new insights into how HDAC inhibitors regulate gene expression. Furthermore, it revealed that the orphaned drug candidate, SR-1815, regulates disease-linked Syngap1 gene expression through direct inhibition of kinases implicated in both neurological disorders and cancer. Through precise mapping of global chromatin mobility, MesoMap promotes insights into how drug-like chemical probes induce transcriptional dynamics within healthy and disease-associated cellular states.

molecular biology↗

bioRxiv2025

The Endo-GeneScreen Platform Identifies Drug-Like Probes that Regulate Endogenous Protein Levels within Physiological Contexts

Traditional phenotypic drug discovery platforms have suffered from poor scalability and a lack of mechanistic understanding of newly discovered phenotypic probes. To address this, we created Endo-GeneScreen (EGS), a high-throughput enabled screening platform that identifies bioactive small molecules capable of regulating endogenous protein expression encoded by any preselected target gene within a biologically appropriate context. As a proof-of-concept, EGS successfully identified drug candidates that up-regulate endogenous expression of neuronal Syngap1, a gene that causes a neurodevelopmental disorder when haploinsufficient. For example, SR-1815, a previously unknown and undescribed kinase inhibitor, alleviated major cellular consequences of Syngap1 loss-of-function by restoring normal SynGAP protein levels and dampening neuronal hyperactivity within haploinsufficient neurons. Moreover, we demonstrate that EGS assays accelerate preclinical development of identified drug candidates and facilitate mode-of-action deconvolution studies. Thus, EGS identifies first-in-class bioactive small molecule probes that promote biological discovery and precision therapeutic development.

neuroscience↗

bioRxiv2025

Proteomic characterisation of the early rheumatoid arthritis-cardiovascular disease multimorbid axis.

BackgroundInflammation contributes to the increased risk of cardiovascular disease (CVD) observed in people with rheumatoid arthritis (RA), with increased prevalence observed from time of diagnosis. Subclinical vascular and myocardial abnormalities can be detected with cardiovascular magnetic resonance (CMR) imaging in otherwise low risk individuals. Identifying associated circulating markers could enable diagnostics and implicate biological pathways of RA-CVD. The studys objective was to identify blood-based proteins associated with CMR measures of vascular and myocardial abnormality and associated inflammatory and/or cardio-metabolic pathways in a new-onset RA cohort. MethodsSerum samples (baseline/pre-treatment, N = 75 and year 1, N = 71) from CADERA (Coronary Artery Disease Evaluation in Rheumatoid Arthritis) participants, a subgroup of a randomised controlled trial, who underwent CMR, were used to measure 334 proteins across 4 pre-defined Olink panels (Inflammation, Cardiovascular-II, Cardiovascular-III, Cardiometabolic). Bayesian mixed effects regression analyses, unsupervised hierarchical clustering and protein network analyses were applied. Normalised protein expression from 334 Olink proteins were used as exposures in the regression analyses. CMR measures of vascular stiffness (aortic distensibility and stiffness index) and myocardial tissue characteristics (native T1, myocardial extracellular volume and late gadolinium enhancement) were each used as individual outcomes in regression analyses. Associations were considered significant at 95% threshold for credible intervals. An expanded physical protein-protein interaction (PPI) network created using the significant (seed) proteins was subjected to topological and enrichment analysis to identify enriched biological pathways. Results54/334 proteins were associated significantly with CMR measures at baseline (7 - vascular; 48 - myocardial tissue characteristics; Coagulation factor 11 with both). Two proteins, TNFSF13B (B-cell activating factor) and CRTAC1, were associated with CMR measures at baseline, were sensitive to change over time and co-varied with changes in CMR measure. Topological analysis of expanded PPI network revealed GRB2 as most connected (degree score=64; closeness score=118.83) and four key signalling pathways, including JAK-STAT and EGFR tyrosine kinase inhibitor resistance, emerged as significant. ConclusionsThis first proteomic study of treatment-naive early RA and subclinical cardiovascular pathology identifies proteins that could aid diagnostic test development and implicates signalling pathways in the RA-cardiovascular axis to inform on our understanding of the basis of RA-CVD.

systems biology↗

bioRxiv2024

Exposure to novel females increases fecundity in adult male prairie voles

Social circuitry of the mammalian brain can influence male reproductive physiology. This often manifests as plasticity in sperm production or allocation, particularly in response to male-male competition. However, socially mediated testicular plasticity has not been investigated with respect to mating and parental strategy. Testis mass and sperm production of sexually naive and female-exposed adult male individuals of three rodent species were compared: the socially monogamous and paternal prairie vole (Microtus ochrogaster), the promiscuous and non-paternal meadow vole (Microtus pennsylvanicus), and the promiscuous and non-paternal house mouse (Mus musculus). Monogamously paired prairie vole males exhibited significantly larger testes and greater sperm production than naive prairie vole males. Comparatively, there were no significant differences between naive and monogamously paired male meadow voles or mice. To investigate the role of olfactory cues for regulating this phenomenon in prairie voles, a group of naive males exposed to soiled bedding from novel females was used. These males were more similar to paired males than to naive males not exposed to novel female odors, demonstrating a strong role of the social olfactory system. Further, while the predictions of sperm competition theory (species with greater female promiscuity have larger testes than closely related species with less female promiscuity) are consistent between naive meadow voles and prairie voles, the prediction does not hold for monogamously paired prairie voles and meadow voles. This demonstrates the complexity of internal social dynamics and reproductive pressures which socially monogamous paternal males face and the evolutionary adaptions that may develop in response.

physiology↗

bioRxiv2024

Development of Clinically Viable Non-Muscle Myosin II Small Molecule Inhibitors with Broad Therapeutic Potential

Non-muscle myosin II (NMII), a molecular motor that regulates critical processes such as cytokinesis and neuronal synaptic plasticity, has substantial therapeutic potential. However, translating this potential to in vivo use has been hampered by the lack of selective tools. The most prototypical non-selective inhibitor, blebbistatin inactivates both NMII and cardiac myosin II (CMII), a key regulator of heart function. Using rational drug design, we developed a series of NMII inhibitors that improve tolerability by selectively targeting NMII over CMII, including MT-228, which has excellent properties such as high brain penetration and efficacy in preclinical models of stimulant use disorder, which has no current FDA-approved therapies. The structure of MT-228 bound to myosin II provides insight into its 17-fold selectivity for NMII over CMII. MT-228s broad therapeutic window opens the door to new disease treatments and provides valuable tools for the scientific community, along with promising leads for future medication development. HighlightsO_LIResearch suggests numerous indications, from axon regeneration and cancer, would benefit from a small molecule inhibitor of non-muscle myosin II, a molecular motor that regulates the actin cytoskeleton. C_LIO_LICurrent chemical probe options are very limited and lack sufficient safety for in vivo C_LIO_LIstudies, which we show is primarily due to potent inhibition of cardiac myosin II. C_LIO_LIRational design that focused on improving target selectivity over the pan-myosin II inhibitor, blebbistatin, led to the identification of MT-228, a small molecule inhibitor with a wide therapeutic window. C_LIO_LIHigh-resolution structure of MT-228 bound to myosin II reveals that selectivity results from a different positioning compared to blebbistatin and an important sequence difference between cardiac and non-muscle myosin II in the inhibitor binding pocket. C_LIO_LIA single administration of MT-228 shows long-lasting efficacy in animal models of stimulant use disorder, a current unmet and rapidly escalating need with no FDA-approved treatments. C_LI

cell biology↗

bioRxiv2024

MT-125 Inhibits Non-Muscle Myosin IIA and IIB, Synergizes with Oncogenic Kinase Inhibitors, and Prolongs Survival in Glioblastoma

We have identified a NMIIA and IIB-specific small molecule inhibitor, MT-125, and have studied its effects in GBM. MT-125 has high brain penetrance and retention and an excellent safety profile; blocks GBM invasion and cytokinesis, consistent with the known roles of NMII; and prolongs survival as a single agent in murine GBM models. MT-125 increases signaling along both the PDGFR- and MAPK-driven pathways through a mechanism that involves the upregulation of reactive oxygen species, and it synergizes with FDA-approved PDGFR and mTOR inhibitors in vitro. Combining MT-125 with sunitinib, a PDGFR inhibitor, or paxalisib, a combined PI3 Kinase/mTOR inhibitor significantly improves survival in orthotopic GBM models over either drug alone, and in the case of sunitinib, markedly prolongs survival in [~]40% of mice. Our results provide a powerful rationale for developing NMII targeting strategies to treat cancer and demonstrate that MT-125 has strong clinical potential for the treatment of GBM. HighlightsO_LIMT-125 is a highly specific small molecule inhibitor of non-muscle myosin IIA and IIB, is well-tolerated, and achieves therapeutic concentrations in the brain with systemic dosing. C_LIO_LITreating preclinical models of glioblastoma with MT-125 produces durable improvements in survival. C_LIO_LIMT-125 stimulates PDGFR- and MAPK-driven signaling in glioblastoma and increases dependency on these pathways. C_LIO_LICombining MT-125 with an FDA-approved PDGFR inhibitor in a mouse GBM model synergizes to improve median survival over either drug alone, and produces tumor free, prolonged survival in over 40% of mice. C_LI

cancer biology↗

bioRxiv2023

Endotoxin, unidentified material, and low amounts of defined toxicants are confounding factors when using the air pollution Standard Reference Material (SRM) 2786 in biological experiments

The National Institute of Standards and Technology (NIST) provides the reference sample SRM 2786 for use in analytical studies of particulate air pollution. The sample originated from an atypical source and possibly contains materials that are unrelated to air pollution. Investigators conducting studies with SRM 2786 should note the relatively low amounts of toxic components listed in the certificate of analysis as well as a large amount of uncharacterized material. Additionally, there is no unsized reference material that can be used as a control for the particulate nature of SRM 2786. The presence of a low amount of endotoxin in SRM 2786 is reported here. Endotoxin and other uncharacterized material in SRM 2786 may influence results and conclusions for biological studies using this reference material.

pharmacology and toxicology↗

bioRxiv2023

Basolateral Amygdala Corticotrophin Releasing Factor Receptor 2 Interacts with Nonmuscle Myosin II to Destabilize Memory

Inhibiting the actin motor ATPase nonmuscle myosin II (NMII) with blebbistatin (Blebb) in the basolateral amgydala (BLA) depolymerizes actin, resulting in an immediate, retrieval-independent disruption of methamphetamine (METH)-associated memory. The effect is highly selective, as NMII inhibition has no effect in other relevant brain regions (e.g. dorsal hippocampus [dPHC], nucleus accumbens [NAc]), nor does it interfere with associations for other aversive or appetitive stimuli, including cocaine (COC). To investigate a potential source of this specificity, pharmacokinetic differences in METH and COC brain exposure were examined. Replicating METHs longer half-life with COC did not render the COC association susceptible to disruption by NMII inhibition. Therefore, transcriptional differences were next assessed. Comparative RNA-seq profiling in the BLA, dHPC and NAc following METH or COC conditioning identified crhr2, which encodes the corticotrophin releasing factor receptor 2 (CRF2), as uniquely upregulated by METH in the BLA. CRF2 antagonism with Astressin-2B (AS2B) had no effect on METH-associated memory after consolidation, allowing for determination of CRF2 influences on NMII-based susceptibility after METH conditioning. Pretreatment with AS2B occluded the ability of Blebb to disrupt an established METH-associated memory. Alternatively, the Blebb-induced, retrieval-independent memory disruption seen with METH was mimicked for COC when combined with CRF2 overexpression in the BLA and its ligand, UCN3 during conditioning. These results indicate that BLA CRF2 receptor activation during learning can prevent stabilization of the actin-myosin cytoskeleton supporting the memory, rendering it vulnerable to disruption via NMII inhibition. CRF2 represents an interesting target for BLA-dependent memory destabilization via downstream effects on NMII.

neuroscience↗

bioRxiv2023

SARS-CoV-2 Mac1 is required for IFN antagonism and efficient virus replication in mice

Several coronavirus (CoV) encoded proteins are being evaluated as targets for antiviral therapies for COVID-19. Included in this set of proteins is the conserved macrodomain, or Mac1, an ADP-ribosylhydrolase and ADP-ribose binding protein. Utilizing point mutant recombinant viruses, Mac1 was shown to be critical for both murine hepatitis virus (MHV) and severe acute respiratory syndrome (SARS)-CoV virulence. However, as a potential drug target, it is imperative to understand how a complete Mac1 deletion impacts the replication and pathogenesis of different CoVs. To this end, we created recombinant bacterial artificial chromosomes (BACs) containing complete Mac1 deletions ({Delta}Mac1) in MHV, MERS-CoV, and SARS-CoV-2. While we were unable to recover infectious virus from MHV or MERS-CoV {Delta}Mac1 BACs, SARS-CoV-2 {Delta}Mac1 was readily recovered from BAC transfection, indicating a stark difference in the requirement for Mac1 between different CoVs. Furthermore, SARS-CoV-2 {Delta}Mac1 replicated at or near wild-type levels in multiple cell lines susceptible to infection. However, in a mouse model of severe infection, {Delta}Mac1 was quickly cleared causing minimal pathology without any morbidity. {Delta}Mac1 SARS-CoV-2 induced increased levels of interferon (IFN) and interferon-stimulated gene (ISG) expression in cell culture and mice, indicating that Mac1 blocks IFN responses which may contribute to its attenuation. {Delta}Mac1 infection also led to a stark reduction in inflammatory monocytes and neutrophils. These results demonstrate that Mac1 only minimally impacts SARS-CoV-2 replication, unlike MHV and MERS-CoV, but is required for SARS-CoV-2 pathogenesis and is a unique antiviral drug target. SIGNIFICANCEAll CoVs, including SARS-CoV-2, encode for a conserved macrodomain (Mac1) that counters host ADP-ribosylation. Prior studies with SARS-CoV-1 and MHV found that Mac1 blocks IFN production and promotes CoV pathogenesis, which has prompted the development of SARS-CoV-2 Mac1 inhibitors. However, development of these compounds into antivirals requires that we understand how SARS-CoV-2 lacking Mac1 replicates and causes disease in vitro and in vivo. Here we found that SARS-CoV-2 containing a complete Mac1 deletion replicates normally in cell culture but induces an elevated IFN response, has reduced viral loads in vivo, and does not cause significant disease in mice. These results will provide a roadmap for testing Mac1 inhibitors, help identify Mac1 functions, and open additional avenues for coronavirus therapies.

microbiology↗

bioRxiv2021

Genomic and transcriptomic somatic alterations of hepatocellular carcinoma in non-cirrhotic livers

BackgroundLiver cancer is the second leading cause of cancer-related deaths worldwide. Hepatocellular carcinoma (HCC) risk factors include chronic hepatitis, cirrhosis, and alcohol abuse, whereby tumorigenesis is induced through inflammation and subsequent fibrotic response. However, a subset of HCC arises in non-cirrhotic livers. We characterized the genomic and transcriptomic landscape of non-cirrhotic HCC to identify features underlying the diseases development and progression. MethodsWhole genome and transcriptome sequencing was performed on 30 surgically resectable tumors comprised of primarily of non-cirrhotic HCC and adjacent normal tissue. Using somatic variants, capture reagents were created and employed on an additional 87 cases of mixed cirrhotic/non-cirrhotic HCC. Cases were analyzed to identify viral integrations, single nucleotide variants (SNVs), insertions and deletions (INDELS), copy number variants, loss of heterozygosity, gene fusions, structural variants, and differential gene expression. ResultsWe detected 3,750 SNVs/INDELS and extensive CNVs and expression changes. Recurrent TERT promoter mutations occurred in >52% of non-cirrhotic discovery samples. Frequently mutated genes included TP53, CTNNB1, and APOB. Cytochrome P450 mediated metabolism was significantly downregulated. Structural variants were observed at MACROD2, WDPCP and NCKAP5 in >20% of samples. Furthermore, NR1H4 fusions involving gene partners EWSR1, GNPTAB, and FNIP1 were detected and validated in 2 non-cirrhotic samples. ConclusionGenomic analysis can elucidate mechanisms that may contribute to non-cirrhotic HCC tumorigenesis. The comparable mutational landscape between cirrhotic and non-cirrhotic HCC supports previous work suggesting a convergence at the genomic level during disease progression. It is therefore possible genomic-based treatments can be applied to both HCC subtypes with progressed disease. HighlightsO_LINon-cirrhotic HCC genomically resembles cirrhotic HCC C_LIO_LIComprehensive genome- and transcriptome-wide profiling allows detection of novel structural variants, fusions, and undiagnosed viral infections C_LIO_LINR1H4 fusions may represent a novel mechanism for tumorigenesis in HCC C_LIO_LINon-cirrhotic HCC is characterized by genotoxic mutational signatures and dysregulated liver metabolism C_LIO_LIClinical history and comprehensive omic profiling incompletely explain underlying etiologies for non-cirrhotic HCC highlighting the need for further research C_LI Short DescriptionThis study characterizes the genomic landscape of hepatocellular carcinomas (HCCs) in non-cirrhotic livers. Using 117 HCCs tumor/normal pairs, we identified 3,750 SNVs/INDELS with high variant frequency in TERT, TP53, CTNNB1, and APOB. CYP450 was significantly downregulated and many structural variants were observed. This characterization could assist in elucidating non-cirrhotic HCC tumorigenesis.

cancer biology↗

bioRxiv2021

Endogenous Syngap1 Alpha Splice Forms Promote Cognitive Function and Seizure Protection

Loss-of-function variants in SYNAGP1 cause a developmental encephalopathy defined by cognitive impairment, autistic features, and epilepsy. SYNGAP1 splicing leads to expression of distinct functional protein isoforms. Splicing imparts multiple cellular functions of SynGAP proteins through coding of distinct C-terminal motifs. However, it remains unknown how these different splice sequences function in vivo to regulate neuronal function and behavior. Reduced expression of SynGAP-1/2 C-terminal splice variants in mice caused severe phenotypes, including reduced survival, impaired learning, and reduced seizure latency. In contrast, upregulation of 1/2 expression improved learning and increased seizure latency. Mice expressing 1-specific mutations, which disrupted SynGAP cellular functions without altering protein expression, promoted seizure, disrupted synapse plasticity, and impaired learning. These findings demonstrate that endogenous SynGAP isoforms with 1/2 spliced sequences promote cognitive function and impart seizure protection. Regulation of SynGAP- expression or function may be a viable therapeutic strategy to broadly improve cognitive function and mitigate seizure.

neuroscience↗

bioRxiv2021

Anatomic development of the upper airway during the first five years of life: A three-dimensional imaging study

PurposeNormative data on the growth and development of the upper airway across the sexes is needed for the diagnosis and treatment of congenital and acquired respiratory anomalies and to gain insight on developmental changes in speech acoustics and disorders with craniofacial anomalies. MethodsThe growth of the upper airway in children ages birth-to-five years, as compared to adults, was quantified using an imaging database with computed tomography studies from typically developing individuals. Methodological criteria for scan inclusion and airway measurements included: head position, histogram-based airway segmentation, anatomic landmark placement, and development of a semi-automatic centerline for data extraction. A comprehensive set of 2D and 3D supra- and sub-glottal measurements from the choanae to tracheal opening were obtained including: naso-oro-laryngo-pharynx subregion volume and length, each subregions superior and inferior cross-sectional-area, and antero-posterior and transverse/width distances. ResultsGrowth of the upper airway during the first five years of life was more pronounced in the vertical and transverse/lateral dimensions than in the antero-posterior dimension. By age five years, females have larger pharyngeal measurement than males. Prepubertal sex-differences were identified in the subglottal region. ConclusionsOur findings demonstrate the importance of studying the growth of the upper airway in 3D. As the lumen length increases, its shape changes, becoming increasingly elliptical during the first five years of life. This study also emphasizes the importance of methodological considerations for both image acquisition and data extraction, as well as the use of consistent anatomic structures in defining pharyngeal regions.

developmental biology↗

bioRxiv2021

Kdm6a Deficiency Restricted to Mouse Hematopoietic Cells Causes an Age- and Sex-dependent Myelodysplastic Syndrome-Like Phenotype

AbstractKdm6a/Utx, a gene on the X chromosome, encodes a histone K27me3 demethylase that has an orthologue on the Y chromosome (Uty)[1]. We previously identified inactivating mutations of Kdm6a in approximately 50% of mouse acute promyelocytic leukemia samples; however, somatic mutations of KDM6A are more rare in human AML samples, ranging in frequency from 2-15% in different series of patients, where their role in pathogenesis is not yet clear. In this study, we show that female Kdm6aflox/flox mice (with allele inactivation initiated by Vav1-Cre in hematopoietic stem and progenitor cells (HSPCs) have a sex-specific phenotype that emerges with aging, with features resembling a myelodysplastic syndrome (MDS). Female Kdm6a-knockout (KO) mice have an age-dependent expansion of their HSPCs with aberrant self-renewal, but they did not differentiate normally into downstream progeny. These mice became mildly anemic and thrombocytopenic, but did not develop overt leukemia, or die from these cytopenias. ChIP-seq and ATAC-seq studies showed only minor changes in H3K27me3, H3K27ac, H3K4me, H3K4me3 and chromatin accessibility between Kdm6a-WT and Kdm6a-KO mice. Utilizing scRNA-seq, Kdm6a loss was linked to the transcriptional repression of genes that mediate hematopoietic cell fate determination. These data demonstrate that Kdm6a plays an important role in normal hematopoiesis, and that its inactivation may contribute to AML pathogenesis by altering the epigenetic state of HSPCs.

genetics↗

bioRxiv2021

Failure to detect mutations U2AF1 due to changes in the GRCh38 reference sequence

The U2AF1 gene is a core part of mRNA splicing machinery and frequently contains somatic mutations that contribute to oncogenesis in MDS, AML, and other cancers. A change introduced in the GRCh38 version of the human reference build prevents mutations in this gene from being detected by many variant calling pipelines. We describe the problem in detail and show that a modified GRCh38 reference build with unchanged coordinates can be used to ameliorate the issue. This reference is available at https://zenodo.org/record/4684553 (doi:10.5281/zenodo.4684553)

genomics↗