Biology subjects

Vlajic, K.

Publications and source records attributed to Vlajic, K..

4 recordsLinked to original sources

bioRxiv2026

Pangenome discovery and characterization of human protein-coding duplicated genes

Protein-coding genes mapping to high-identity segmental duplications (SDs) have been difficult to annotate and characterize and are the source of most previously unknown protein-coding genes being discovered as part of the human pangenome. Here, we combine long-read assembled human genomes (298) and long-read transcriptome data (5.6 billion full-length cDNA from 83 tissues) to phylogenetically interrogate 493 gene families discovering 2713 potentially copy number polymorphic genes not present in the human reference genome. For reference SD gene families where paralog specificity can be assigned, we find that 60.0% are expressed and maintain open reading frames, with 45.7% showing high expression in brain, embryo, or testis. We revise 386 gene models, including 150 that absent or different from current T2T-CHM13 gene annotation and 236 (35.1%) pseudogenes as protein-coding where we find evidence of transcription, an open reading frame, and chromatin-accessible promoters. We find that 24.2% of SD genes show evidence of constraint for both copy number and amino acid mutation. The majority of these constraint genes are ancestral, whereas only 16.2% of derived duplicated genes that emerged recently in the human lineage show evidence of constraint. The pangenome provides unparalleled specificity to understand genetic variation in SD genes allowing us to distinguish functional genes from pseudogenes and highlighting potential gene innovations that arose most recently in human evolution.

genomics↗

bioRxiv2026

Detergent selection as a determinant of lysate melting behavior and drug-target hit-calling in PISA

Thermal proteome profiling (TPP) and proteome integral solubility alteration (PISA) assays measure drug-target interactions by monitoring protein thermal stability across the proteome. While detergents are routinely used in lysate-based thermal profiling, the field lacks consensus on whether detergents should be present during the melting step or only added afterward as an extraction buffer, and whether detergent identity matters for this choice. Here, we evaluate how commonly used detergents and the timing of their use in thermal stability workflows affect proteome-wide thermal stability and PISA hit calling in TF-1 lysates. We find that NP-40 and DDM produce highly correlated melting profiles when used exclusively as post-melt extraction buffers, but diverge substantially when present during the melting step. DDM in particular prevents the thermally-induced loss in solubility of large classes of proteins, such as cell surface proteins, and these effects propagate directly into PISA hit calling. Performing the PISA melt in DDM versus NP-40 results in the gain and loss of distinct drug-target interactions for both the PAK4 inhibitor PF-3758309 and the PLK1 inhibitor volasertib. Notably, DDM enables detection of a volasertib-TMEM97 interaction that was previously not detected in NP-40. However, we also find that the stabilization effects of DDM mask the identification of some known PISA hits for these drugs. We further introduce a four-parameter logistic model of protein melting to aid in modeling of these findings and a linear regression framework for PISA hit calling that outperforms pairwise t-tests in low-replicate settings. Together, these results establish detergent selection as a tunable experimental variable in thermal profiling and suggest that some drug-target engagements previously attributed exclusively to intact-cell context may be recoverable in lysates with appropriate buffer conditions.

biochemistry↗

bioRxiv2026

Multimodal analysis of molecular remodeling in aging spleen identified global and cell type specific changes

Aging reshapes the cellular and molecular landscape of mammalian tissues. These changes can be progressive, preceding linearly with age, or occur as abrupt transitions of the course of lifespan. To investigate the age-dependent cellular and molecular shifts we profiled matched proteomes and transcriptomes from male and female murine spleens across eight time points, from stable adults through late life. The spleen was chosen to integrate understanding of age-dependent changes associated with immune surveillance, inflammaging, and immune-related proteostasis. Male and female mice follow distinct aging trajectories particularly in protein-RNA correlation in late life, reflecting both compositional shifts and failure of post-transcriptional buffering. To investigate whether these changes could be attributed to specific cell-types within the spleen, we developed Celestial, a machine-learning framework to identify cell-type-specific changes in bulk tissue samples. We found that age-related bulk molecular changes could be attributed in part to compositional remodeling of cell-types--expansion of GZMK+ CD8+ T cells and C1Q+ macrophages alongside naive T cell and global B cell loss. These results demonstrate that cell-type-aware interpretation can inform bulk multi-omic data for accurate mechanistic inference in heterogeneous tissues undergoing complex molecular remodeling.

systems biology↗

bioRxiv2026

Serum proteomics reveals distinct phenotypic signatures to IL-6 blockade between two immunotherapies

A recent clinical study tested the effects of two different monoclonal antibodies (mAbs) (siltuximab, anti-IL6; tocilizumab, anti-IL6R) on the fate and function of T-cells in people with type 1 diabetes. While both mAbs affect the response of T-cells to stimulation, they have very different, sometimes opposing mechanisms. Here, we use mass-spectrometry based proteomics to analyze longitudinal serum samples (baseline and two weeks post-treatment) from 20 clinical trial participants to examine the effects of siltuximab and tocilizumab on extracellular vesicles. To accomplish this, serum samples were enriched for extracellular vesicles with Mag-Net and analyzed by LC-MS/MS to identify significantly differentially abundant protein groups and pathways. Proteome analysis confirmed highly reproducible measurements across multiple draw dates. In total, we quantified >3300 protein groups of which 46 protein groups had significantly altered abundance after mAb treatment. Tocilizumab altered pathways associated with proteostasis (neddylation) and pre-notch transcription and translation. Siltuximab altered FCGR activation pathway members. In addition, quantitation of the monoclonal antibody therapies themselves enabled the measurement of the correlation between drug amounts and impacted proteins. Taken together, this work demonstrates the utility of the Mag-Net method to evaluate the impacts of therapeutic interventions on serum extracellular vesicles.

biochemistry↗