Biology subjects

Nava, A. A.

Publications and source records attributed to Nava, A. A..

5 recordsLinked to original sources

bioRxiv2023

KAT6A mutations in Arboleda-Tham syndrome drive epigenetic regulation of posterior HOXC cluster

Arboleda-Tham Syndrome (ARTHS) is a rare genetic disorder caused by heterozygous, de novo truncating mutations in Lysine(K) acetyltransferase 6A (KAT6A). ARTHS is clinically heterogeneous and characterized by several common features including intellectual disability, developmental and speech delay, hypotonia and affects multiple organ systems. KAT6A is highly expressed in early development and plays a key role in cell-type specific differentiation. KAT6A is the enzymatic core of a histone-acetylation protein complex, however the direct histone targets and gene regulatory effects remain unknown. In this study, we use ARTHS patient (n=8) and control (n=14) dermal fibroblasts and perform comprehensive profiling of the epigenome and transcriptome caused by KAT6A mutations. We identified differential chromatin accessibility within the promoter or gene body of 23%(14/60) of genes that were differentially expressed between ARTHS and controls. Within fibroblasts, we show a distinct set of genes from the posterior HOXC gene cluster (HOXC10, HOXC11, HOXC-AS3, HOXC-AS2, HOTAIR) that are overexpressed in ARTHS and are transcription factors critical for early development body segment patterning. The genomic loci harboring HOXC genes are epigenetically regulated with increased chromatin accessibility, high levels of H3K23ac, and increased gene-body DNA methylation compared to controls, all of which are consistent with transcriptomic overexpression. Finally, we used unbiased proteomic mass spectrometry and identified two new histone post-translational modifications (PTMs) that are disrupted in ARTHS: H2A and H3K56 acetylation. Our multi-omics assays have identified novel histone and gene regulatory roles of KAT6A in a large group of ARTHS patients harboring diverse pathogenic mutations. This work provides insight into the role of KAT6A on the epigenomic regulation in somatic cell types.

genomics↗

bioRxiv2023

Characterization and diversification of AraC/XylS family regulators guided by transposon sequencing

In this study, we explored the development of engineered inducible systems. Publicly available data from previous transposon sequencing assays were used to identify regulators of metabolism in Pseudomonas putida KT2440. For the AraC-family regulators (AFRs) represented in this data, we posited AFR/promoter/inducer groupings. Eleven promoters were characterized for a response to their proposed inducers in P. putida, and the resultant data were used to create and test nine two-plasmid sensor systems in E. coli. Several of these were further developed into a palette of single-plasmid inducible systems. From these experiments, we observed an unreported inducer response from a previously characterized AFR, demonstrated that the addition of a P. putida transporter improved the sensor dynamics of an AFR in E. coli, and identified an uncharacterized AFR with a novel potential inducer specificity. Finally, targeted mutations in an AFR, informed by structural predictions, enabled further diversification of these inducible plasmids. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=105 SRC="FIGDIR/small/550116v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@fb1ee2org.highwire.dtl.DTLVardef@a88f1dorg.highwire.dtl.DTLVardef@4bbd5borg.highwire.dtl.DTLVardef@7cdbfc_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2023

KAT6A mutations drive transcriptional dysregulation of cell cycle and Autism risk genes in an Arboleda-Tham Syndrome cerebral organoid model

Arboleda-Tham Syndrome (ARTHS, OMIM#616268) is a rare neurodevelopmental disorder caused by de novo mutations in KAT6A. Individuals with ARTHS typically exhibit varying degrees of intellectual disability, speech and language deficits and clinical manifestations across multiple systems that lead to abnormal: vision, craniofacial features, cardiac morphology, and gastrointestinal function. To gain insight into the potential neuropathological mechanisms underlying ARTHS, we investigate how KAT6A mutations disrupt in vitro brain development using induced pluripotent stem cells (iPSCs) and cerebral organoids (COs) derived from ARTHS patients harboring KAT6A nonsense mutations. In this study, we conducted comprehensive transcriptomic profiling by performing time-course experiments and generating short-read and long-read RNA sequencing (RNA-seq) data from undifferentiated iPSCs and COs at 15 and 25 days of neural differentiation. Our analysis revealed abnormal expression of 235 genes in ARTHS across all three timepoints examined. Notably, we observed persistent dysregulation of genes such as CTSF, ZNF229, PCDHB12, and PAK3. Additionally, we found a consistent enrichment of PTBP1-target genes among the upregulated genes in ARTHS at all three stages assessed by RNA-seq. During neural differentiation, we identified 980 genes that consistently display aberrant transcription in ARTHS at both CO stages. These genes are enriched for genes involved in cell fate determination through modulation of cell-cycle dynamics (e.g. E2F family) and cell-adhesion molecules (e.g. PCDH genes). Our findings indicate that ARTHS COs exhibit slower downregulation of pluripotency and cell cycle genes compared to controls and that this delay led to an overrepresentation of cycling human neural progenitor markers during neural differentiation in ARTHS. Finally, matching the variable neurodevelopment phenotypes in ARTHS, we discovered that the aberrantly expressed genes in ARTHS are enriched for genes associated with Autism Spectrum Disorder and Epilepsy, with a subset showing isoform-specific dysregulation. Strikingly, the same PTBP1-target genes were enriched amongst the genes that display differential isoform usage in ARTHS. For the first time, we demonstrate that KAT6A mutations lead to a delay in repressing pluripotency and cell cycle genes during neural differentiation, suggesting that prolonged activation of these gene networks disrupts the temporal dynamics of human brain development in ARTHS.

genetics↗

bioRxiv2023

Maximizing Heterologous Expression of Engineered Type I Polyketide Synthases: Investigating Codon Optimization Strategies

Type I polyketide synthases (T1PKSs) hold an enormous potential as a rational production platform for the biosynthesis of specialty chemicals. However, despite the great progress in this field, the heterologous expression of PKSs remains a major challenge. One of the first measures to improve heterologous gene expression can be codon optimization. Although controversial, choosing the wrong codon optimization strategy can have detrimental effects on protein and product levels. In this study, we analyzed 11 different codon variants of an engineered T1PKS and investigated in a systematic approach their influence on heterologous expression in Corynebacterium glutamicum, Escherichia coli, and Pseudomonas putida. Our best performing codon variants exhibited a minimum 50-fold increase in PKS protein levels, which also enables the production of an unnatural polyketide in each of the hosts. Furthermore, we developed a free online tool (https://basebuddy.lbl.gov) that offers transparent and highly customizable codon optimization with up-to-date codon usage tables. Here, we not only highlight the significance of codon optimization but also establish the groundwork for high-throughput assembly and characterization of PKS pathways in alternative hosts.

synthetic biology↗

bioRxiv2023

Foldy: a web application for interactive protein structure analysis

Foldy is a cloud-based application that allows non-expert scientists to easily access and utilize advanced AI-based structural biology tools, including AlphaFold and DiffDock. Built on Kubernetes, it can be deployed by universities, departments, and labs without requiring hardware resources, but can also be configured to utilize available computers. Foldy enables scientists to predict the structure of proteins and complexes up to 3000 amino acids, visualize Pfam annotations, and dock ligands with AutoDock Vina and DiffDock. Our manuscript describes the user interface and deployment considerations of Foldy, as well as some of our applications. By democratizing access to sophisticated AI-based tools, Foldy can facilitate life science research and promote the wider adoption of structural bioinformatics tools. Our work demonstrates that even the most advanced tools can be made accessible to a broad audience through user-friendly platforms like Foldy, and we believe it will be a valuable resource for researchers across scientific disciplines. The public structures available on the Lawrence Berkeley Labs Foldy deployment can be viewed at https://foldy.lbl.gov. Author SummaryFoldy is a cloud-based application that enables scientists to use AI-based structural biology tools such as AlphaFold and DiffDock without software expertise. Built on Kubernetes, it can be set up by universities, departments, and labs with no need for hardware resources. Foldy can predict the structure of proteins and complexes up to 3000 amino acids, visualize Pfam annotations, and dock ligands with AutoDock Vina and DiffDock. Our public structures can be viewed at https://foldy.lbl.gov. Our manuscript highlights the user interface, deployment considerations, and product applications of Foldy. Its an accessible solution for researchers who are not software experts and can handle the traffic of thousands of users and hundreds of thousands of protein structures and docked ligands. This makes advanced AI-based tools more widely available, paving the way for accelerating life science research. By developing an easy-to-use platform, our work demonstrates that even the most sophisticated AI-based tools can be made accessible to a wide audience. Foldy enables more scientists to draw from the rapidly growing field of structural biology, making it a valuable tool for researchers across scientific disciplines. We look forward to its adoption by the scientific community.

bioinformatics↗