Biology subjects

Malek, M.

Publications and source records attributed to Malek, M..

10 recordsLinked to original sources

bioRxiv2026

Spatially resolved tumor-microenvironment architecture defines therapeutic response in unresectable hepatoblastoma

Background: Hepatoblastoma exhibits variable therapeutic response that is not fully explained by current risk stratification. While prior studies have characterized tumor composition, the role of spatial organization in determining outcome, particularly in unresectable disease requiring transplantation under immunosuppression, remains unclear. Methods: We integrated spatial transcriptomics using the 10x Genomics Visium platform (n = 11 tumors) with single-cell spatial molecular imaging using NanoString CosMx (n = 3 tumors) on post-chemotherapy explant specimens across clinically defined groups (relapse-prone unresectable, non-relapse unresectable, and resectable hepatoblastoma). Spatial deconvolution, module scoring, spatial co-localization, and cell-cell interaction analyses were used to define region-level transcriptional programs and their underlying cellular organization. Results: We identified three spatial architectures associated with clinical outcome. Relapse-prone tumors exhibited a centralized, TME-dominated organization enriched for progenitor-like, cycling, and stress-responsive tumor cell states co-localized with SPP1+ TAMs and CAFs, with enrichment of hypoxia response, extracellular matrix remodeling, and Midkine (MDK-LRP1) signaling. Non-relapse tumors displayed a distributed architecture with compartmentalized tumor-immune interactions, increased immune engagement, and preserved epithelial differentiation, including inflammatory hepatocyte-like states. Resection samples showed reduced tumor cell state heterogeneity and more uniform TME-immune organization. Conclusions: Integration of regional and single-cell spatial transcriptomics across two platforms reveals that the chemo-remodeled tumor architecture of hepatoblastoma encodes spatially organized tumor cell-TME interactions as central determinants of therapeutic response.

cancer biology↗

bioRxiv2026

Structural Connectivity Between the Zinc Linchpin Motif, the cluster, and the active site orchestrates DNA repair in MUTYH

The DNA glycosylase MUTYH protects genomic integrity by excising adenine mispaired with 8-oxoguanine (OG), which initiates base excision repair (BER). The [4Fe-4S] cluster DNA binding domain and Zn linchpin motif of MUTYH are hotspots for inherited cancer associated variants (CAVs) highlighting their critical functions in DNA repair. Here, we present three full-length human MUTYH crystal structures bound to DNA across three catalytic states, representing early and late transition states and product complexes. These structures reveal a previously unrecognized interplay between the Zn site and the Fe-S cluster mediated by His85 and a conserved Arg247/Arg307 bridge spanning [~]15 [A]. Disruption of this network impairs metal loading, reduces the active enzyme fraction, weakens lesion DNA binding, and diminishes OG:A repair in cells. Moreover, the Zn site adopts distinct coordination modes across the reaction coordinate, with a water molecule replacing a cysteine ligand in transition state analog complexes and cysteine coordination restored in a product-mimicked state. Functional analyses show that this ligand switching is dispensable for core glycosylase chemistry in vitro but disproportionately affects repair in cells, suggesting an additional role of the Zn linchpin in cellular OG:A repair beyond intrinsic glycosylase activity, likely involving interactions with BER partners. Together, these findings illustrate how the Zn linchpin enhances Fe-S cluster domain DNA substrate engagement to enable effective DNA repair and provides a rationale for how these functions are compromised by MUTYH CAVs.

biochemistry↗

bioRxiv2026

MUTYH cancer-associated variants within the interdomain connector differentially impact glycosylase activity and cellular DNA repair

The base excision repair (BER) glycosylase MUTYH initiates repair of 8-oxo-7,8-dihydroguanine (OG): adenine (A) mispairs to prevent G to T transversion mutations. Inherited biallelic mutations in MUTYH are correlated with the cancer pre-disposition syndrome MUTYH-associated polyposis (MAP) and contribute to an increased lifetime risk of colorectal cancer. Over 1,000 germline and somatic MUTYH variants have been reported that are associated with MAP and other cancers, but for most the functional impact is unknown. Herein, we examined a subset of cancer-associated variants (CAVs) localized in the interdomain connector (IDC), which links the N-terminal adenine excision and C-terminal OG recognition domains via its zinc linchpin motif and serves as a hub for downstream repair interactions. In vitro assays measuring glycosylase activity, lesion affinity, and AP endonuclease stimulation revealed no substantial defects relative to wild-type MUTYH. In contrast, a newly optimized mammalian cell assay revealed some IDC variants exhibit reduced repair. These results suggest that some variants disrupt steps downstream of adenine excision, whereas others impair lesion recognition and base excision. This work underscores the value of independent functional assays for accurately assessing variant dysfunction and classification. Analysis of MUTYH variants highlights the complexity of the roles of MUTYH in preserving genomic integrity.

biochemistry↗

bioRxiv2025

Citizen science gamers enable automated flow cytometry gating through machine learning

Manual flow cytometry gating requires up to one hour per sample with 32% inter-expert variability, creating critical bottlenecks in immunological research reproducibility. To address this, we developed flowMagic, a machine learning algorithm for automated gating that is trained on both expert-curated data (template data) and crowdsourced annotations from citizen science gaming. Through EVE Online, 839,199 players analyzed 52,178 bivariate plots from 37 studies, generating 31,703 quality-controlled training plots. Evaluated against 92,203 expert-validated files spanning 79 immune populations (i.e., a biologically defined cell subset within each bivariate plot), flowMagic achieved 90% accuracy for abundant populations and 65% for rare populations, outperforming existing methods. The algorithm reproduced biological patterns including neutrophil dynamics in COVID-19 patients and immune development in newborns. This gaming-based approach demonstrates that crowd-sourced pattern recognition generates robust training data for complex biomedical applications, offering transformative potential for standardizing flow cytometry analysis and accelerating immunological discovery.

bioinformatics↗

bioRxiv2025

Crystal structure of MutYX: A novel clusterless adenine DNA glycosylase with a distinct C-terminal domain and 8-Oxoguanine recognition sphere

The [4Fe-4S] cluster is an important cofactor of the base excision repair (BER) adenine DNA glycosylase MutY to prevent mutations associated with 8-oxoguanine (OG). Several MutYs lacking the [4Fe-4S] cofactor have been identified. Phylogenetic analysis shows that clusterless MutYs are distributed in two clades suggesting cofactor loss in two independent evolutionary events. Herein, we determined the first crystal structure of a clusterless MutY complexed with DNA. On the basis of the dramatic structural divergence from canonical MutYs, we refer to this as representative of a clusterless MutY subgroup "MutYX". Interestingly, MutYX compensates for the missing [4Fe-4S] cofactor to maintain positioning of catalytic residues by expanding a pre-existing -helix and acquisition of the new -helix. Surprisingly, MutYX also acquired a new C-terminal domain that uniquely recognizes OG using residue Gln201 and Arg209. Adenine glycosylase assays and binding affinity measurements indicate that Arg209 is the primary residue responsible to specificity for OG:A lesions, while Gln201 bridges OG and Arg209. Surprisingly, replacement of Arg209 and Gln201 with Ala increases activity toward G:A mismatches. The MutYX structure serves as an example of devolution, capturing structural features required to retain function in the absence of a metal cofactor considered indispensable.

biochemistry↗

bioRxiv2023

Spatial transcriptome profiling uncovers metabolic regulation of left-right patterning

Left-right patterning disturbance can cause severe birth defects, but it remains least understood of the three body axes. We uncovered an unexpected role for metabolic regulation in left-right patterning. Analysis of the first spatial transcriptome profile of left-right patterning revealed global activation of glycolysis, accompanied by right-sided expression of Bmp7 and genes regulating insulin growth factor signaling. Cardiomyocyte differentiation was left-biased, which may underlie the specification of heart looping orientation. This is consistent with known Bmp7 stimulation of glycolysis and glycolysis suppression of cardiomyocyte differentiation. Liver/lung laterality may be specified via similar metabolic regulation of endoderm differentiation. Myo1d, found to be left-sided, was shown to regulate gut looping in mice, zebrafish, and human. Together these findings indicate metabolic regulation of left-right patterning. This could underlie high incidence of heterotaxy-related birth defects in maternal diabetes, and the association of PFKP, allosteric enzyme regulating glycolysis, with heterotaxy. This transcriptome dataset will be invaluable for interrogating birth defects involving laterality disturbance.

developmental biology↗

bioRxiv2023

Differential Expression of Gluconeogenesis-Related Transcripts in a Freshwater Zooplankton Model Organism Suggests a Role of the Cori Cycle in Hypoxia Tolerance

O_LIGluconeogenesis (GNG) is the process of regenerating glucose and NAD+ that allows continuing ATP synthesis by glycolysis during fasting or in hypoxia. Recent data from C. elegans and crustaceans challenged with hypoxia show differential and tissue-specific expression of GNG-specific genes. C_LIO_LIHere we report differential expression of several GNG-specific genes in the head and body of a model organism, Daphnia magna, a planktonic crustacean, in normoxic and acute hypoxic conditions. We predict that GNG-specific transcripts will be enriched in the body, where most of the fat tissue is located, rather than in the head, where the tissues critical for survival in hypoxia, the central nervous system and locomotory muscles, are located. We measured the relative expression of GNG-specific transcripts in each body part by qRT-PCR and normalized them by either the expression of a reference gene or the rate-limiting glycolysis enzyme pyruvate kinase (PK). C_LIO_LIOur data show that of the three GNG-specific transcripts tested, pyruvate carboxylase (PC) showed no differential expression in either the head or body. Phosphoenolpyruvate carboxykinase (PEPCK-C), on the other hand, is upregulated in hypoxia in both body parts. Fructose-1,6-bisphosphatase (FBP) is upregulated in the body relative to the head and upregulated in hypoxia relative to normoxia, with a stronger body effect in hypoxia when normalized by PK expression. C_LIO_LIThese results support our hypothesis that Daphnia can survive hypoxic conditions by implementing the Cori cycle, where body tissues supply glucose and NAD+ to the brain and muscles, enabling them to continuously generate ATP by glycolysis. C_LI

biochemistry↗

bioRxiv2022

Prior selection affects phenotypic and transcriptional response to hypoxia

Hypoxia has profound and diverse effects on aerobic organisms, disrupting oxidative phosphorylation and activating several protective pathways. Predictions have been made that exposure to mild intermittent hypoxia may be protective against more severe exposure and may extend lifespan. Both effects are likely to depend on prior selection on phenotypic and transcriptional plasticity in response to hypoxia, and may therefore show signs of local adaptation. Here we report the lifespan effects of chronic, mild, intermittent hypoxia (CMIH) and short-term survival in acute severe hypoxia (ASH) in four clones of Daphnia magna originating from either permanent or intermittent habitats, the latter regularly drying up with frequent hypoxic conditions. We show that CMIH extended the lifespan in the two clones originating from intermittent habitats but had the opposite effect in the two clones from permanent habitats, which also showed lower tolerance to ASH. Exposure to CMIH did not protect against ASH; to the contrary, Daphnia from the CMIH treatment had lower ASH tolerance than normoxic controls. Few transcripts changed their abundance in response to the CMIH treatment in any of the clones. After 12 hours of ASH treatment, the transcriptional response was more pronounced, with numerous protein-coding genes with functionality in mitochondrial and respiratory metabolism, oxygen transport, and gluconeogenesis showing up-regulation. While clones from intermittent habitats showed somewhat stronger differential expression in response to ASH than those from permanent habitats, there were no significant hypoxia-by-habitat of origin or CMIH-by-ASH interactions. GO enrichment analysis revealed a possible hypoxia tolerance role by accelerating the molting cycle and regulating neuron survival through up-regulation of cuticular proteins and neurotrophins, respectively.

ecology↗

bioRxiv2020

Inference and validation of an integrated regulatory network of autism

Autism is a complex neurodevelopmental disorder. Functional roles of several non-coding transcripts including long noncoding RNAs (lncRNAs) have been shown to influence the pathobiology of autism. We hypothesized that there are more autism-associated lncRNAs to be discovered. Here, we utilized a systems biology approach to identify novel lncRNAs that might play a role in the molecular pathogenesis of autism. Based on the data provided by the Simons Foundation Autism Research Initiative (SFARI), a three-component regulatory network comprising mRNAs, microRNAs (miRNAs) and lncRNAs was constructed. Functional enrichment analysis was performed to identify molecular pathways potentially mediated by components of the network. The potential association of four candidate lncRNAs with autism was investigated experimentally by developing and verifying a valproic acid (VPA)-exposed mouse model of autism. We composed a network of 33 mRNA, 25 miRNA and 4 lncRNA nodes associated with neurologically-relevant pathways and functions. We then verified the differential expression of four candidate lncRNAs: Gm10033, 1500011B03Rik, A930005H10Rik and Gas5 in the brain of VPA-exposed mice. We furthermore identified a novel splice variant of Gm10033, designated as Gm10033-{Delta}Ex2, which was expressed in various mouse tissues. The integrative approach, we utilized, combines the analysis of a three-component regulatory network with experimental validation of targets in an animal model of autism. As a result of the analysis, we prioritized a set of candidate autism-associated lncRNAs. These links add to the common understanding of the molecular and cellular mechanisms that are involved in disease etiology, specifically in the autism.

systems biology↗

bioRxiv2018

Descriptors of Pain Quality Associate with Pulpal and Periapical Diagnostic Tests in Patients Experiencing Acute Tooth Pain

Introduction Introduction Material And Methods Results Discussion References Dental pain causes a great deal of suffering, stress, and interference in the lives of patients, and relief from pain is a common reason for seeking dental care (Heaivilin et al. 2011). However, how we typically measure pain in dentistry does little to capture the complex experience of pain. Scales such as the visual analog scale (VAS) or numeric rating scale (NRS) are the most commonly used, and allow for the quantification of pain intensity. Used alone, as is most often then case, these scales are quite limited, given the multidimensional nature of pain, with sensory, affective, and temporal dimensions mediated by complex and varied biological mechanisms (Dworkin et al. 2005; von Hehn et al. 2012). Pain ...

neuroscience↗