E2F3 is a central regulator of cell cycle progression, development, differentiation, and oncogenesis. Alternative promoter usage generates two major homologous isoforms, E2F3A and E2F3B, differing only at their N-termini. Although E2F3A is traditionally considered an activator and E2F3B a repressor of transcription, studies suggest that both isoforms have overlapping and isoform-specific functions, shaped by differences in temporal expression, protein stability, and target gene specificity. In this review, we examine how E2F3A and E2F3B contribute to proliferation, differentiation, and cancer. We also discuss contradictions and limitations in experimental approaches that have complicated the interpretation of these functions. Finally, we explore how clarifying these roles could help resolve long-standing questions in cell cycle regulation, differentiation, and cancer, where E2F3 plays a major role.

Abstract. Activity-based profiling with fluorescent probes is a powerful tool for the functional characterization of whole enzyme classes in crude proteome

CONCLUSION: These findings indicate that the ELISA assays tested, designed to measure specific isoforms of mature CGRP isoforms, are unable to detect the synthetic peptide. Nevertheless, it is possible that these assays quantify other CALCA/CALCB-derived products that may reta...

In a transcriptome, isoforms share most of their sequence, and gene families add further near-copies. Yet most long-read mappers treat each isoform as a separate target and repeat seeding, chaining an...

AbstractAKT protein kinases are vital serine/threonine kinases that function as central regulators of the PI3K/AKT signaling pathway, which controls various biological processes, including cell survival, growth, proliferation, and metabolism. Among the three AKT isoforms, AKT3 remains the least characterized, despite its involvement in brain malformations and cancers. Post-translational modifications (PTMs), particularly phosphorylation, regulate AKT function and activity, yet the proteoform landscape of AKT3 has not been characterized. Here, we performed top-down Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) to characterize full-length recombinant human AKT3. Ultrahigh-resolution intact mass analysis resolved AKT3 proteoforms carrying up to three phosphorylations, with unphosphorylated and monophosphorylated species accounting for 40% and 39% of the total proteoform abundance, respectively. Complementary collisionally activated dissociation (CAD) and electron-capture dissociation (ECD) confirmed the removal of the initiating methionine and localized the N-terminal acetylation. Targeted tandem mass spectrometry (MS/MS) of monophosphorylated AKT3 generated terminal and internal fragments that localized the primary phosphorylation site to Thr332 (canonical Thr305). These findings uncover AKT3 proteoform diversity and demonstrate the power of ultrahigh-resolution top-down FTICR-MS for resolving and characterizing intact kinase proteoforms.

Read the full paper: Context-dependent toxicity of human tau isoforms in a Drosophila tauopathy model

Mutations in the microtubule-binding protein tau are considered causal for a diverse group of neurodegenerative diseases collectively referred to as primary tauopathies. Specific tauopathies are associated with different mutations, distinct tau isoforms, manifest in unique cellular and circuit-wide patterns of degeneration, and present with specific clinical symptoms and trajectories. The mechanisms that link variations in the primary tau protein structure to distinct forms of neural circuit dysfunction remain largely unknown. Here, we created an orthogonal viral resource to systematically examine the impact of expressing humanized tau isoforms harboring different disease-relevant mutations in excitatory pyramidal cells from the mouse prelimbic cortex. We used these reagents to show that expression of mutant humanized tau (hTau) as either of two isoforms produced very similar patterns of circuit-specific synaptic disconnection on cortical projection neurons. However, mass spectrometry revealed that the interactomes of these different tau species varied markedly as functions of isoform, mutation, and subcellular location. We validated the functional relevance of increased hTau interactions with mitochondrial proteins in induced neurons differentiated from Ngn2-iPSCs expressing different hTau constructs. These results suggest different disease-relevant hTau proteins likely use distinct mechanisms to drive a highly stereotyped form of synapse loss in cortical neurons.