Granular component sub-phases direct ribosome biogenesis in the nucleolus.

The hierarchical, multiphase organization of the nucleolus underlies ribosome biogenesis. Ribonucleoprotein particles that regulate ribosomal subunit assembly are heterogeneously disposed in the granular component (GC) of the nucleolus. However, the molecular origins of the GC's spatial heterogeneity and its association with ribosomal subunit assembly remain poorly understood.

Assessing Protein Surface-Based Scoring for Interpreting Genomic Variants.

Clinical genomics sequencing is rapidly expanding the number of variants that need to be functionally elucidated. Interpreting genetic variants (i.e.

An Image Analysis Pipeline for Quantifying the Features of Fluorescently-Labeled Biomolecular Condensates in Cells.

Biomolecular condensates are cellular organelles formed through liquid-liquid phase separation (LLPS) that play critical roles in cellular functions including signaling, transcription, translation, and stress response. Importantly, condensate misregulation is associated with human diseases, including neurodegeneration and cancer among others. When condensate-forming biomolecules are fluorescently-labeled and examined with fluorescence microscopy they appear as illuminated foci, or puncta, in cells.

Enhanced interpretation of 935 hotspot and non-hotspot RAS variants using evidence-based structural bioinformatics.

In the current study, we report computational scores for advancing genomic interpretation of disease-associated genomic variation in members of the RAS family of genes. For this purpose, we applied 31 sequence- and 3D structure-based computational scores, chosen by their breadth of biophysical properties. We parametrized our data by assembling a numerically homogenized experimentally-derived dataset, which when use in our calculations reveal that computational scores using 3D structure highly correlate with experimental measures (e.

Phase Separation Mediates NUP98 Fusion Oncoprotein Leukemic Transformation.

Unlabelled: NUP98 fusion oncoproteins (FO) are drivers in pediatric leukemias and many transform hematopoietic cells. Most NUP98 FOs harbor an intrinsically disordered region from NUP98 that is prone to liquid-liquid phase separation (LLPS) in vitro. A predominant class of NUP98 FOs, including NUP98-HOXA9 (NHA9), retains a DNA-binding homeodomain, whereas others harbor other types of DNA- or chromatin-binding domains.

Computational modeling reveals key molecular properties and dynamic behavior of disruptor of telomeric silencing 1-like (DOT1L) and partnering complexes involved in leukemogenesis.

Disruptor of telomeric silencing 1-like (DOT1L) is the only non-SET domain histone lysine methyltransferase (KMT) and writer of H3K79 methylation on nucleosomes marked by H2B ubiquitination. DOT1L has elicited significant attention because of its interaction or fusion with members of the AF protein family in blood cell biology and leukemogenic transformation. Here, our goal was to extend previous structural information by performing a robust molecular dynamic study of DOT1L and its leukemogenic partners combined with mutational analysis.

Molecular mechanics and dynamic simulations of well-known Kabuki syndrome-associated KDM6A variants reveal putative mechanisms of dysfunction.

Background: Kabuki syndrome is a genetic disorder that affects several body systems and presents with variations in symptoms and severity. The syndrome is named for a common phenotype of faces resembling stage makeup used in a Japanese traditional theatrical art named kabuki. The most frequent cause of this syndrome is mutations in the H3K4 family of histone methyltransferases while a smaller percentage results from genetic alterations affecting the histone demethylase, KDM6A.

Structural bioinformatics enhances mechanistic interpretation of genomic variation, demonstrated through the analyses of 935 distinct RAS family mutations.

Motivation: Protein-coding genetic alterations are frequently observed in Clinical Genetics, but the high yield of variants of uncertain significance remains a limitation in decision making. RAS-family GTPases are cancer drivers, but only 54 variants, across all family members, fall within well-known hotspots. However, extensive sequencing has identified 881 non-hotspot variants for which significance remains to be investigated.

Novel destabilizing Dynactin variant (DCTN1 p.Tyr78His) in patient with Perry syndrome.

Introduction: Perry syndrome, also recognized as Perry disease, is a rare autosomal dominant disorder characterized by midlife-onset atypical parkinsonism, apathy or depression, respiratory failure and weight loss caused by a mutation in the Dynactin (DCTN1) gene.

Case Description: A fifty-six years-old adopted male presented with atypical parkinsonism with bradykinesia and postural instability, apathy, weight loss, and recurrent respiratory failure due to central hypoventilation requiring tracheostomy.

Methods And Results: Clinical workup revealed a novel DCTN1 p.

Opposing Intermolecular Tuning of Ca Affinity for Calmodulin by Neurogranin and CaMKII Peptides.

We investigated the impact of bound calmodulin (CaM)-target compound structure on the affinity of calcium (Ca) by integrating coarse-grained models and all-atomistic simulations with nonequilibrium physics. We focused on binding between CaM and two specific targets, Ca/CaM-dependent protein kinase II (CaMKII) and neurogranin (Ng), as they both regulate CaM-dependent Ca signaling pathways in neurons. It was shown experimentally that Ca/CaM (holoCaM) binds to the CaMKII peptide with overwhelmingly higher affinity than Ca-free CaM (apoCaM); the binding of CaMKII peptide to CaM in return increases the Ca affinity for CaM.

Correlation between Gene Variants, Signaling Pathways, and Efficacy of Chemotherapy Drugs against Colon Cancers.

Efficacies, toxicities, and resistance mechanisms of chemotherapy drugs, such as oxaliplatin and 5-fluorouracil (5-FU), vary widely among various categories and subcategories of colon cancers. By understanding the differences in the drug efficacy and resistance at the level of protein-protein networks, we identified the correlation between the drug activity of oxaliplatin/5-FU and gene variations from the US National Cancer Institute-60 human cancer cell lines. The activity of either of these drugs is correlated with specific amino acid variant(s) of KRAS and other genes from the signaling pathways of colon cancer progression.

Absence of Activation of DNA Repair Genes and Excellent Efficacy of Phosphaplatins against Human Ovarian Cancers: Implications To Treat Resistant Cancers.

Phosphaplatins, platinum(II) and platinum(IV) complexes coordinated to a pyrophosphate moiety, exhibit excellent antitumor activities against a variety of cancers. To determine whether phosphaplatins trigger resistance to treatment by engaging DNA damage repair genes, a yeast genome-wide fitness assay was used. Treatment of yeast cells with pyrodach-2 (D2) or pyrodach-4 (D4) revealed no particular sensitivity to nucleotide excision repair, homologous recombination repair, or postreplication repair when compared with platin control compounds.

Alterations of Nonconserved Residues Affect Protein Stability and Folding Dynamics through Charge-Charge Interactions.

Charge-charge interactions play an important role in thermal stability of proteins. We employed an all-atom, native-topology-based model with non-native electrostatics to explore the interplay between folding dynamics and stability of TNfn3 (the third fibronectin type III domain from tenascin-C). Our study elucidates the role of charge-charge interactions in modulating the folding energy landscape.

Lessons in Protein Design from Combined Evolution and Conformational Dynamics.

Protein-protein interactions play important roles in the control of every cellular process. How natural selection has optimized protein design to produce molecules capable of binding to many partner proteins is a fascinating problem but not well understood. Here, we performed a combinatorial analysis of protein sequence evolution and conformational dynamics to study how calmodulin (CaM), which plays essential roles in calcium signaling pathways, has adapted to bind to a large number of partner proteins.

Conformational frustration in calmodulin-target recognition.

Calmodulin (CaM) is a primary calcium (Ca(2+) )-signaling protein that specifically recognizes and activates highly diverse target proteins. We explored the molecular basis of target recognition of CaM with peptides representing the CaM-binding domains from two Ca(2+) -CaM-dependent kinases, CaMKI and CaMKII, by employing experimentally constrained molecular simulations. Detailed binding route analysis revealed that the two CaM target peptides, although similar in length and net charge, follow distinct routes that lead to a higher binding frustration in the CaM-CaMKII complex than in the CaM-CaMKI complex.

Allostery and folding of the N-terminal receiver domain of protein NtrC.

The N-terminal receiver domain of protein NtrC (NtrC(r)) exhibits allosteric transitions between the inactive (unphosphorylated) and active (phosphorylated) state on the microsecond time scale. Using a coarse-grained variational model with coupled energy basins, we illustrate that significant loss of conformational flexibility is the key determinant of the inactive (I) → active (A) state transition mechanism of NtrC(r). In particular, our results reveal that the rearrangements of the native contacts involving the regulatory helix-α4 and the flexible β3-α3 loop upon activation play a crucial role in the activation mechanism.

Backtracking due to residual structure in the unfolded state changes the folding of the third fibronectin type III domain from tenascin-C.

Residual structure in the unfolded state of a protein may play a crucial role in folding and stability. In the present study, using an all (heavy)-atom structure based model and replica exchange molecular dynamics simulations, we explored the folding landscape of the third fibronectin type III domain from tenascin-C (TNfn3). Specifically, both the wild type (WT) and a variant with two additional amino acids, Gly-Leu (GL), at the C-terminus (WT(+GL)) were studied.

Conformational flexibility and the mechanisms of allosteric transitions in topologically similar proteins.

Conformational flexibility plays a central role in allosteric transition of proteins. In this paper, we extend the analysis of our previous study [S. Tripathi and J.