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Biology subjects

Palma, A.

Publications and source records attributed to Palma, A..

At least 19 recordsLinked to original sources

Generative inverse design steers human hematopoietic cell fate in vitro

Inverse design has advanced the discovery of molecules and materials. Yet it remains largely unexplored for steering cell fate, where inputs are continuous, combinatorial culture conditions and outputs are distributions of cell states. Human hematopoiesis, in which combinations and doses of growth factors steer stem cells toward each blood lineage, is a demanding test case. While forward models can evaluate discrete conditions, the vast combinatorial space renders searching for a specific, target-producing design an intractable inverse problem. Here, we introduce LabCompass, an inverse-modeling framework that optimizes experimental designs toward candidates expected to produce specified target cell states, using a flow-matching generative model guided by cellular phenotypes predicted by a forward surrogate. We applied LabCompass to primary human CD34+ in vitro cultures, where continuous cytokine and small-molecule parameters govern hematopoietic cell fate. In a lab-in-the-loop setting, we experimentally validated model-driven designs and used these measurements to update the model across four successive loops. LabCompass recovered established lineage-inducing designs, improved predictions with new data, and enriched target populations across lineages. Incorporating new parameters expanded the design space and enriched rare populations beyond initial reach. Across loops, we generated a 78-million-cell spectral flow cytometry dataset and identified distinct culture conditions converging on the same target cell states. Our results establish generative inverse modeling as a compelling paradigm for navigating high-dimensional, continuous experimental spaces, and demonstrate that virtual cell models should be judged not only by how accurately they predict cellular responses, but also by how effectively they guide experiments toward desired biological outcomes.

cell biology↗

Benchmarking gene expression reconstruction from single-cell latent representations

Single-cell transcriptomics is typically modeled in low-dimensional latent representations that improve the signal-to-noise ratio of the data. Such representations underpin data integration, cell state discovery, and perturbation prediction, with applications ranging from large-scale organ atlases to latent trajectory modeling. Recent virtual cell approaches further leverage these representations to predict cellular responses as distributional shifts in latent space. Each of these applications ultimately requires faithful gene expression reconstruction from latent spaces for biological interpretation, enabling gene-level analysis of predicted perturbed or batch-corrected cells. Yet representation choice is typically treated as an implementation detail rather than a primary modeling decision, with no systematic evaluation of how well latent representations support gene expression reconstruction. Here, we introduce ReconEval, a benchmark for evaluating gene expression reconstruction from single-cell latent spaces. We benchmark two classes of latent representations: end-to-end trained models such as PCA, autoencoders, and variational autoencoders, and pretrained single-cell foundation model embeddings coupled to newly trained decoders. Reconstruction is evaluated both directly and after latent-space perturbation prediction. Across perturbational and observational datasets totaling over 100 million cells, our metric suite quantifies statistical fidelity; biological signal preservation, including differential expression, coexpression, cell-cycle structure, cytokine response and pathway activity; and perturbation-specific effects. We find that autoencoders achieve the strongest stand-alone reconstruction at low dimensionality, while variational regularization does not improve generalization in reconstruction. Frozen foundation model embeddings retain recoverable gene-level information, with reconstruction quality depending strongly on decoder architecture and pretraining objective. In latent perturbation modeling, high-dimensional PCA matches foundation model embeddings, while low-dimensional AE embeddings are optimal for flow-based generative models. Overall, reconstruction depends critically on the interplay between representation and downstream model, and simpler representations can outperform complex alternatives given appropriate capacity. Our benchmark establishes reconstruction as a critical evaluation axis for single-cell foundation models. We envision it improving the biological interpretability of latent-space modeling, a prerequisite for future virtual cell models to be validated by domain experts and grounded in biology.

bioinformatics↗

A systematic approach to understanding how patient variants affect the activity of Wiskott-Aldrich syndrome protein

Wiskott-Aldrich syndrome (WAS) and X-linked neutropenia (XLN) are caused by genetic variants in the WAS gene. How WAS variants lead to clinical disease remains unsolved in many cases. We expressed human WASp using a spider silk inspired solubility tag (NT*-tag) and inserted patients variants. Native mass spectrometry and pyrene actin assays showed that five variants (L270P, F271S, S272P, I290T, I294T) predicted to cause XLN led to open protein conformation and high actin polymerization rate in the absence of the WASp activator, Cdc42. One previously reported XLN variant (R268W), two loss-of-function WAS variants (A236G, D485N), and one variant of unknown significance (R431W) behaved similarly to wildtype WASp in terms of structural conformation and actin polymerization. Patient CD4+ T cells were used for analysis of WASp expression and phosphorylation, actin polymerization, anti-CD3 induced proliferation capacity, and upregulation of high affinity LFA-1, distinguishing loss-of-function and gain-of-function variants from benign WAS variants. This systematic approach reveals how WAS genetic variants cause severe human disease and stratify variants to guide clinical decision for definitive therapy. Key PointsO_LIGain-of-function WASp variant has extended protein conformation probed by native mass spectrometry and raised pyrene actin polymerization. C_LIO_LIFunctional analysis of patients CD4+ T cells classifies WASp variants as loss-of-function, reduced-function, gain-of-function, and benign. C_LI

immunology↗

Ribosome Quality Control Mitigates Proteotoxic Stress in Aneuploid Cells

Aneuploidy is widespread in tumors, but how cancer cells adapt to aneuploidy-induced cellular stresses remains poorly understood. Here, we focus on the mechanisms employed to cope with proteostasis disruption, a major stress caused by aneuploidy. We show that aneuploid cells exhibit a significant accumulation of ribosomes enclosed within autophagosomes, ultimately degraded through lysosome-mediated processes. Our data also indicate that limited folding capacity of newly synthesized polypeptides leads to lysosome-mediated degradation of ribosomes. We also found that the E3 ligase ZNF598 marks these ribosomes for degradation, thus clearing translationally-impaired ribosomes. Importantly, highly aneuploid tumors display a positive correlation with ZNF598 expression, while being negatively associated with ribosomal signatures. This suggests that ribosome-associated quality control is crucial for cancer cell survival under proteotoxic stress. Our study uncovers molecular events in response to proteotoxic stress in aneuploid cells and suggests that components of ribosome-associated quality control, including ZNF598, could serve as promising targets in cancer therapy.

cell biology↗

scPortrait integrates single-cell images into multimodal modeling

Machine learning increasingly uncovers rules of biology directly from data, enabled by large, standardized datasets. Microscopy images provide rich information on cellular architecture and are accessible at scale across biological systems, making them an ideal foundation for modeling cell behavior. However, a standardized image format does not exist at the single-cell level. Here we present scPortrait, an scverse software package for generation, storage, and application of single-cell image datasets. scPortrait reads, stitches and segments raw fields of view with out-of-core computation scaling to larger-than-memory datasets. Parallelization enables rapid extraction of individual cells into a standardized single-cell image format with fast access to accelerate machine learning. scPortrait enables analysis across modalities including images, proteomics and transcriptomics, identifying cancer-associated macrophage subpopulations by morphology and embedding single-cell images into transcriptome atlases. scPortrait turns microscopy images into a reusable resource for integrative cell modeling, establishing single-cell images as a core modality in systems biology.

systems biology↗

CellFlow enables generative single-cell phenotype modeling with flow matching

High-content phenotypic screens provide a powerful strategy for studying biological systems, but the scale of possible perturbations and cell states makes exhaustive experiments unfeasible. Computational models that are trained on existing data and extrapolate to correctly predict outcomes in unseen contexts have the potential to accelerate biological discovery. Here, we present CellFlow, a flexible framework based on flow matching that can model single cell phenotypes induced by complex perturbations. We apply CellFlow to various phenotypic screens, accurately predicting expression responses to a wide range of perturbations, including cytokine stimulation, drug treatments and gene knockouts. CellFlow successfully modeled developmental perturbations at the whole-embryo scale and guided cell fate and organoid engineering by predicting heterogeneous cell populations arising from combinatorial morphogen treatments and by performing a virtual organoid protocol screen. Taken together, CellFlow has the potential to accelerate discovery from phenotypic screens by learning from existing data and generating phenotypes induced by unseen conditions.

bioinformatics↗

Long noncoding RNA HSCHARME promotes the maturation of stem cells-derived cardiomyocytes by controlling gene regulatory networks altered in human cardiomyopathies

A growing body of evidence suggests that tissue-specific long noncoding RNAs (lncRNA) play pivotal roles in the heart. Here, we exploited the synteny between the mouse and human genomes to identify the novel lncRNA HSCHARME (Human Syntenic CHARME) and combined single-cell transcriptomics, CAGE-seq data, RNA-FISH imaging and CRISPR-Cas9 genome editing to document its role in cardiomyogenesis. By investigating the mechanism of action of HSCHARME in hiPSC-derived cardiomyocytes, we found that the locus produces the major pCHARME isoform that associates with SC35-containing speckles and interacts with the splicing regulator PTBP1. Consistently, the functional inactivation of pCHARME influences the splicing of cardiac-specific pre-mRNAs and impacts their expression, which parallels a decline in cardiomyocyte differentiation and physiology. In line with a possible association with disease, large-scale analysis of the lncRNA expression across cardiomyopathy patients revealed increased levels of pCHARME in hypertrophic (HCM) and dilated (DCM) hearts and identified a subset of disease-associated targets whose expression can be modulated through HSCHARME dosage. By unlocking mechanistic insights into the role of pCHARME in cardiac cells, our data identify a novel non-coding regulator of cardiomyocyte function with potential implications in disease.

molecular biology↗

A circuit involving the lncRNA MB3 and the driver genes MYC and OTX2 inhibits apoptosis in Group 3 Medulloblastoma by regulating the TGF-β pathway via HMGN5

BackgroundGroup 3 (G3) is one of the most common, aggressive and fatal subtypes of the paediatric cerebellar tumour Medulloblastoma (MB), primarily driven by the MYC oncogene. Targeting MYC has long been challenging and this, combined with our incomplete understanding of G3 MB molecular bases, has hindered the development of effective targeted therapies. Long noncoding RNAs (lncRNAs), with their extensive oncogenic roles, cancer-specific expression, and connection to MYC biology, offer opportunities for unravelling this complexity and providing new insights and therapeutic targets. MethodologyUsing genome-wide, molecular and cellular assays, we characterised the activity of the MYC-dependent, anti-apoptotic lncRNA lncMB3 in G3 MB cells. ResultsThrough transcriptomic and interactomic analyses, we clarified lncMB3 function and mode-of-action. LncMB3 controls the TGF-{beta} pathway, critically altered in G3 medulloblastomagenesis. This regulation occurs via the direct coding-noncoding RNA interaction between lncMB3 and the mRNA for the epigenetic factor HMGN5, with both sharing targets in the TGF-{beta} cascade. This axis converges on apoptosis through OTX2, another G3 MB driver gene, and photoreceptor lineage genes. Synergistic effects between lncMB3 targeting and cisplatin treatment underscores the relevance of this regulatory network in vitro. Finally, we propose novel ferritin-based nanocarriers as efficient delivery tools for antisense oligonucleotides targeting lncMB3. ConclusionsLncMB3 emerges as a central node linking MYC amplification to apoptosis inhibition through a circuit involving RNA-based mechanisms, G3 MB key drivers and underexplored factors. This integrated framework deepens our understanding of G3 MB molecular underpinnings and lay the foundation for translating lncRNA research into potential applications.

cancer biology↗

Genome biology of long non-coding RNAs in humans: a virtual karyotype

BackgroundLong non-coding RNAs (lncRNAs) represent a unique and groundbreaking class of RNA molecules that exert regulatory functions with remarkable tissue and cellular specificities. Although the number of identified functional lncRNAs is increasing, comprehensive profiling of lncRNA genomics remains elusive. Creating a virtual lncRNA karyotype is especially important for species whose intrinsic features enable their biosynthesis and function in context-dependent manners. Results and conclusionsTo address this challenge, we employed existing annotation files to create a statistical genomics portrait of lncRNA genes for comparison with protein-coding genes. We provide a foundational reference for exploring the non-coding genome, offering insights into the genomic characteristics of lncRNAs that may enhance understanding of their biological significance and impact.

genomics↗

Unveiling microglial heterogeneity from single-cell transcriptomics in neurodegenerative diseases

Microglia are key players in maintaining brain homeostasis and responding to pathological conditions. Their multifaceted roles in health and disease have garnered significant attention in the context of neurodegeneration. In recent years, single-cell transcriptomic techniques have provided unprecedented insights into microglial heterogeneity, revealing distinct subpopulations and gene expression patterns associated with neuroprotection or neurotoxicity. Here, the transcriptomic landscape of microglia has been dissected by leveraging human single-nuclei RNA sequencing datasets of neurodegenerative conditions, encompassing amyotrophic lateral sclerosis, frontotemporal dementia, Alzheimers disease, aging, and Parkinsons disease. Results have led to the identification of distinct cell subpopulations, representative of the functional heterogeneity of the brain microglia. Moreover, distinct gene signatures and regulatory networks linked to inflammation and neurodegeneration have been identified. Overall, the study provides an improved portrait of microglia in the context of neurodegenerative disorders, and it holds promise for developing a more targeted research aimed at modulating microglial function to mitigate disease progression and foster neuroprotection. HighlightsO_LIBrain-resident microglia cells show a profound transcriptional heterogeneity C_LIO_LIMicroglia are not subject to a macrophage-like polarization mechanism C_LIO_LINeurodegenerative disorders share common transcriptional programs, yet retaining their own peculiarities C_LIO_LIDistinct gene regulatory networks underlie microglia heterogeneity and neurodegeneration-related dysfunctions C_LI

neuroscience↗

Reexamining the essentiality of Pdi1 in yeast: A PDI1 knockout is viable in Komagataella phaffii and still produces recombinant disulfide bonded proteins

Protein disulfide isomerase 1 (Pdi1) and Ero1 form the main oxidative folding axis in the endoplasmic reticulum (ER). Despite having additional PDI family members, Pdi1 has long ago been determined to be essential in bakers yeast and this concept was inferred to other yeast species. In this study, we reexamine the gene essentiality of Pdi1 in the methylotrophic yeast Komagataella phaffii. Strikingly, the absence of Pdi1 does not cause lethality, but even allows for folding and secretion of heterologous proteins in a homogeneous redox state. Remarkably, while sensitivity to externally added folding stressors, such as tunicamycin and DTT is increased in pdi1{Delta} knock-out cells, in non-stressed growth conditions they do not display upregulation of folding stress markers, such as the master chaperone Kar2 or the unfolded protein response regulator Hac1, suggesting homeostatic adaptation.

microbiology↗

Transcriptional profiling of human brain cortex identifies novel lncRNA-mediated networks dysregulated in amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease for which a comprehensive knowledge about the pathological mechanisms is still lacking. A multitude of dysregulated cellular processes and pathways have been linked to ALS so far, including the recent focus directed toward the implication of several classes of non-coding (nc)RNAs. Within this context, the class of long ncRNAs (lncRNAs), may provide an important contribution to the onset and the severity of ALS pathogenesis, due to their high tissue specificity and their function as gene expression regulators. Nevertheless, their identification in humans often relies on differential expression analyses from bulk RNA-seq, which limits their targeting in the cellular contexts where they may be primarily involved. Here we apply dedicated pipelines to single-nucleus nuclei datasets to study lncRNA from non-pathological and pre-frontal ALS human cortex. We found that in brain, distinct cell subtypes express very different pattern of lncRNAs to suggest possible roles in cellular processes found dysregulated in ALS patients. Moreover, we show the lncRNA involvement in important gene regulatory networks that result differentially regulated in pathological conditions and dissect the genomic organization of differentially expressed lncRNAs.

genomics↗

SC2Spa: a deep learning based approach to map transcriptome to spatial origins at cellular resolution

BackgroundUnderstanding cellular heterogeneity within tissues hinges on knowledge of their spatial context. However, it is still challenging to accurately map cells to their spatial coordinates. ResultsWe present SC2Spa, a deep learning-based approach that learns intricate spatial relationships from spatial transcriptomics (ST) data. Benchmarking tests show that SC2Spa outperformed other predictors and accurately detected tissue architecture from transcriptome. SC2Spa successfully mapped single cell RNA sequencing (scRNA-seq) to Visium assay, providing an approach to enhance the resolution for low resolution ST data. Our test showed that SC2Spa performs well for various ST technologies and robust to spatial resolution. In addition, SC2Spa can suggest spatially variable genes that cannot be identified from previous approaches. ConclusionsSC2Spa is a robust and accurate approach to provide single cells with their spatial location and identify spatially meaningful genes.

bioinformatics↗

Predicting cell morphological responses to perturbations using generative modeling

Advancements in high-throughput screening have enabled the exploration of rich phenotypic readouts like high-content microscopy, expediting drug target identification and mode of action studies. However, scaling these experiments to the vast space of drug or genetic manipulations poses challenges, as only a small subset of compounds show activity in screenings. Despite being widely used in various applications, machine learning methods have not shown a reliable ability to extrapolate predictions to scenarios involving unseen phenomena, specifically transforming an unseen control cell image into a desired perturbation. We present a generative model, the IMage Perturbation Autoencoder (IMPA), which predicts cellular morphological effects of chemical and genetic perturbations using untreated cells as input. IMPA learns perturbation-specific styles from generalized embeddings and generates counterfactual treatment response predictions in control cells. We demonstrate IMPA can predict morphological changes caused by small molecule perturbations on breast cancer cells. Additionally, we test IMPA on the unseen drug effect prediction task, showing improved performance over state-of-the-art generative models when compounds are structurally related to the training set. Finally, generalizability and capability to predict more subtle effects are showcased through its application to large microscopy datasets with hundreds of genetic perturbations on U2OS cells. We envision IMPA to become a valuable tool in computational microscopy for aiding phenotypic drug discovery, facilitating navigation of the perturbation space, and rational experimental design.

bioinformatics↗

CHK2 Regulates MUS81-EME2 Function in Response to Replication Stress and BRCA2 Deficiency

MUS81 is a structure-specific endonuclease that processes DNA intermediates in mitosis and in S-phase following replication stress. MUS81 is crucial to cleave deprotected reversed forks in BRCA2-deficient cells. However, how MUS81 is regulated during replication stress in human cells remains unknown. Our study reveals that CHK2 binds the MUS81-EME2 complex and positively regulates formation of DSBs upon replication stress or in the absence of BRCA2. The association with MUS81 occurs through the FHA domain of CHK2 and is disabled by the I157T mutation but not by the R117A mutation. The CHK2-MUS81 complex forms downstream fork reversal and degradation, and phosphorylation of MUS81 at CHK2-targeted sites is crucial to introduce DSBs at deprotected replication forks ensuring the replication fork recovery in BRCA2-deficient cells. Collectively, our work sheds light into the regulation of the MUS81 complex and identifies a novel function of the ATM-CHK2 axis in the response to deprotected replication forks in the absence of BRCA2.

cell biology↗

A new insight into MYC action: control of RNA polymerase II methylation and transcription termination

A common catastrophic event in most human cancers is deregulation of MYC, a multifunctional transcription factor that controls gene expression in partnership with MAX and drives key biological mechanisms of the cell. Restraining its activity impairs cancer cell features and prevents tumor development, as shown by Omomyc - a 90 amino acid mini-protein interfering with MYC activity. MYC regulates many aspects of transcription by RNA polymerase II (RNAPII), such as activation, pause release, and elongation. That it may have a role in transcription termination as well is suggested by our finding of an interaction between MYC and the Protein Arginine Methyltransferase 5 (PRMT5), which catalyzes symmetrical dimethylation of RNAPII at the arginine residue R1810 (R1810me2s) allowing proper termination and splicing of transcripts. Here we show that MYC overexpression strongly increases R1810me2s, while the concomitant expression of Omomyc or a MYC-specific shRNA counteracts this capacity. Omomyc impairs as well Serine 2 phosphorylation in the RNAPII carboxyterminal domain, a modification that sustains transcript elongation and is enhanced by MYC. By displacing MYC on DNA, Omomyc reshapes RNAPII distribution along genes, leading to greater occupancy of promoter and termination sites. It is unclear how this may affect expression of the variety of genes that control metabolic, biosynthetic, and other pathways and are up or down regulated upon MYC inhibition. Genes belonging to a signature of direct MYC targets are instead strongly downregulated following MYC inhibition, with a weak correlation with RNAPII occupancy at promoters. Our data point to a MYC/ PRMT5/RNAPII axis that controls termination via RNAPII dimethylation (R1810me2s) and may contribute to fine-tune the expression of genes altered by MYC overexpression in cancer cells. It remains to be seen which role this may have in tumor development and maintenance.

cancer biology↗

Clemastine induces an impairment in developmental myelination

Abnormalities in myelination are associated to behavioral and cognitive dysfunction in neurodevelopmental psychiatric disorders. Thus, therapies to promote or accelerate myelination could potentially ameliorate symptoms in autism. Clemastine, a histamine H1 antagonist with anticholinergic properties against muscarinic M1 receptor, is the most promising drug with promyelinating properties (Mei et al., 2014). Clemastine penetrates the blood brain barrier efficiently and promotes remyelination in different animal models of neurodegeneration including multiple sclerosis, ischemia and Alzheimers disease. However, its role in myelination during development is unknown. We showed that clemastine treatment during development increase oligodendrocyte differentiation in both white and gray matter. However, despite the increase in the number of oligodendrocytes, conduction velocity of myelinated fibers of corpus callosum decreased in clemastine treated mice. Confocal and electron microscopy showed a reduction in the number of myelinated axons and nodes of Ranvier and a reduction of myelin thickness in corpus callosum. To understand the mechanisms leading to myelin formation impairment in the presence of an excess of myelinating oligodendrocytes, we focused on microglial cells which also express muscarinic M1 receptors. Importantly, the population of CD11c+microglia cells, necessary for myelination, as well as the levels of insulin growth factor-1 decrease in clemastine-treated mice. Altogether, these data suggest that clemastine impact on myelin development is more complex than previously thought and could be dependent on microglia-oligodendrocyte crosstalk. Further studies are needed to clarify the role of microglia cells on developmental myelination.

neuroscience↗

Substantial Differences in SARS-CoV-2 Antibody Responses Elicited by Natural Infection and mRNA Vaccination

We analyzed data from two ongoing COVID-19 longitudinal serological surveys in Orange County, CA., between April 2020 and March 2021. A total of 8,476 finger stick blood specimens were collected before and after an aggressive mRNA vaccination campaign. IgG levels were determined using a multiplex antigen microarray containing 10 SARS-CoV-2 antigens, 4 SARS, 3 MERS, 12 Common CoV, and 8 Influenza antigens. Twenty-six percent of 3,347 specimens from unvaccinated Orange County residents in December 2020 were SARS-CoV-2 seropositive. The Ab response was predominantly against nucleocapsid (NP), full length spike and the spike S2 domain. Anti-receptor binding domain (RBD) reactivity was low and there was no cross-reactivity against SARS S1 or SARS RBD. An aggressive mRNA vaccination campaign at the UCI Medical Center started on December 16, 2020 and 6,724 healthcare workers were vaccinated within 3 weeks. Seroprevalence increased from 13% in December to 79% in January, 93% in February and 99% in March. mRNA vaccination induced much higher Ab levels especially against the RBD domain and significant cross-reactivity against SARS RBD and S1 was also observed. Nucleocapsid protein Abs can be used to distinguish individuals in a population of vaccinees to classify those who have been previously infected and those who have not, because nucleocapsid is not in the vaccine. Previously infected individuals developed higher Ab titers to the vaccine than those who have not been previously exposed. These results indicate that mRNA vaccination rapidly induces a much stronger and broader Ab response than SARS-CoV-2 infection.

immunology↗