Atypical thymomas with squamoid and spindle cell features: clinicopathologic, immunohistochemical and molecular genetic study of 120 cases with long-term follow-up.

Thymomas are rare tumors characterized by a broad range of morphologic appearances that can sometimes give rise to difficulties for classification. We have studied a series of 120 thymoma patients in whom the tumors were characterized by sheets of atypical epithelial cells with squamoid and/or spindle cell features. They occurred in 63 men and 57 women and presented as a discrete mass in the anterior mediastinum measuring 2-23 cm (mean: 8.2 cm). Patients' ages ranged from 14 to 86 years (mean: 57.8) and most had symptoms referable to a mass lesion. 20 patients had myasthenia gravis or other autoimmune disorder. 76 cases were characterized by a predominant population of round to polygonal tumor cells while 32 cases were characterized by atypical oval or spindle cells. 12 cases showed mixed features and 16 cases showed the development of thymic carcinoma arising from thymoma. All cases were positive for p40/p63 and cytokeratin AE1/AE3. 23 cases were positive for CD5 (25%), and 13 for CD117 (14%). MIB1 showed a significant increase in proliferative activity (mean = 11.6%). Next generation sequencing in 47 cases did not disclose any variants amenable to current targeted therapies. Clinical follow up ranging from 2 to 29 years showed a progressive increase in aggressive behavior and fatality rate with advancing stage. Overall survival was 87% at 5 years, 67% at 10 years, and 23% at 20 years. Completeness of resection and staging were the most significant parameters for survival. The more aggressive tumors followed a protracted clinical course with multiple recurrences and metastases over a long period of time (mean = 19.8 years from time of initial relapse to death). Atypical thymomas are a distinct category of thymic epithelial neoplasm characterized by a slowly progressive clinical course with increased potential for metastases, transformation to a higher-grade malignancy, and fatal outcome in some cases.

Fine-tuned repression of Drp1-driven mitochondrial fission primes a 'stem/progenitor-like state' to support neoplastic transformation.

Gene knockout of the master regulator of mitochondrial fission, Drp1, prevents neoplastic transformation. Also, mitochondrial fission and its opposing process of mitochondrial fusion are emerging as crucial regulators of stemness. Intriguingly, stem/progenitor cells maintaining repressed mitochondrial fission are primed for self-renewal and proliferation. Using our newly derived carcinogen transformed human cell model, we demonstrate that fine-tuned Drp1 repression primes a slow cycling 'stem/progenitor-like state', which is characterized by small networks of fused mitochondria and a gene-expression profile with elevated functional stem/progenitor markers (Krt15, Sox2 etc) and their regulators (Cyclin E). Fine tuning Drp1 protein by reducing its activating phosphorylation sustains the neoplastic stem/progenitor cell markers. Whereas, fine-tuned reduction of Drp1 protein maintains the characteristic mitochondrial shape and gene-expression of the primed 'stem/progenitor-like state' to accelerate neoplastic transformation, and more complete reduction of Drp1 protein prevents it. Therefore, our data highlights a 'goldilocks' level of Drp1 repression supporting stem/progenitor state dependent neoplastic transformation.

New quantitative approach reveals heterogeneity in mitochondrial structure-function relations in tumor-initiating cells.

Steady-state mitochondrial structure or morphology is primarily maintained by a balance of opposing fission and fusion events between individual mitochondria, which is collectively referred to as mitochondrial dynamics. The details of the bidirectional relationship between the status of mitochondrial dynamics (structure) and energetics (function) require methods to integrate these mitochondrial aspects. To study the quantitative relationship between the status of mitochondrial dynamics (fission, fusion, matrix continuity and diameter) and energetics (ATP and redox), we have developed an analytical approach called mito-SinCe2 After validating and providing proof of principle, we applied mito-SinCe2 on ovarian tumor-initiating cells (ovTICs). Mito-SinCe2 analyses led to the hypothesis that mitochondria-dependent ovTICs interconvert between three states, that have distinct relationships between mitochondrial energetics and dynamics. Interestingly, fusion and ATP increase linearly with each other only once a certain level of fusion is attained. Moreover, mitochondrial dynamics status changes linearly with ATP or with redox, but not simultaneously with both. Furthermore, mito-SinCe2 analyses can potentially predict new quantitative features of the opposing fission versus fusion relationship and classify cells into functional classes based on their mito-SinCe2 states.This article has an associated First Person interview with the first author of the paper.

Crosstalk between the mitochondrial fission protein, Drp1, and the cell cycle is identified across various cancer types and can impact survival of epithelial ovarian cancer patients.

Mitochondrial metabolic reprogramming is a hallmark of tumorigenesis. Although mitochondrial function can impact cell cycle regulation it has been an understudied area in cancer research. Our study highlights a specific involvement of mitochondria in cell cycle regulation across cancer types. The mitochondrial fission process, which is regulated at the core by Drp1, impacts various cellular functions. Drp1 has been implicated in various cancer types with no common mechanism reported. Our Drp1-directed large-scale analyses of the publically available cancer genomes reveal a robust correlation of Drp1 with cell-cycle genes in 29 of the 31 cancer types examined. Hypothesis driven investigation on epithelial ovarian cancer (EOC) revealed that Drp1 co-expresses specifically with the cell-cycle module responsible for mitotic transition. Repression of Drp1 in EOC cells can specifically attenuate mitotic transition, establishing a potential casual role of Drp1 in mitotic transition. Interestingly, Drp1-Cell-Cycle co-expression module is specifically detected in primary epithelial ovarian tumors that robustly responded to chemotherapy, suggesting that Drp1 driven mitosis may underlie chemo-sensitivity of the primary tumors. Analyses of matched primary and relapsed EOC samples revealed a Drp1-based-gene-expression-signature that could identify patients with poor survival probabilities from their primary tumors. Our results imply that around 60% of platinum-sensitive EOC patients undergoing relapse show poor survival, potentially due to further activation of a mitochondria driven cell-cycle regime in their recurrent disease. We speculate that this patient group could possibly benefit from mitochondria directed therapies that are being currently evaluated at various levels, thus enabling targeted or personalized therapy based cancer management.

A new mitochondrial pool of cyclin E, regulated by Drp1, is linked to cell-density-dependent cell proliferation.

The regulation and function of the crucial cell cycle regulator cyclin E (CycE) remains elusive. Unlike other cyclins, CycE can be uniquely controlled by mitochondrial energetics, the exact mechanism being unclear. Using mammalian cells (in vitro) and Drosophila (in vivo) model systems in parallel, we show that CycE can be directly regulated by mitochondria through its recruitment to the organelle. Active mitochondrial bioenergetics maintains a distinct mitochondrial pool of CycE (mtCycE) lacking a key phosphorylation required for its degradation. Loss of the mitochondrial fission protein dynamin-related protein 1 (Drp1, SwissProt O00429 in humans) augments mitochondrial respiration and elevates the mtCycE pool allowing CycE deregulation, cell cycle alterations and enrichment of stem cell markers. Such CycE deregulation after Drp1 loss attenuates cell proliferation in low-cell-density environments. However, in high-cell-density environments, elevated MEK-ERK signaling in the absence of Drp1 releases mtCycE to support escape of contact inhibition and maintain aberrant cell proliferation. Such Drp1-driven regulation of CycE recruitment to mitochondria might be a mechanism to modulate CycE degradation during normal developmental processes as well as in tumorigenic events.

Biological systems discovery in silico: radical S-adenosylmethionine protein families and their target peptides for posttranslational modification.

Data mining methods in bioinformatics and comparative genomics commonly rely on working definitions of protein families from prior computation. Partial phylogenetic profiling (PPP), by contrast, optimizes family sizes during its searches for the cooccurring protein families that serve different roles in the same biological system. In a large-scale investigation of the incredibly diverse radical S-adenosylmethionine (SAM) enzyme superfamily, PPP aided in building a collection of 68 TIGRFAMs hidden Markov models (HMMs) that define nonoverlapping and functionally distinct subfamilies. Many identify radical SAM enzymes as molecular markers for multicomponent biological systems; HMMs defining their partner proteins also were constructed. Newly found systems include five groupings of protein families in which at least one marker is a radical SAM enzyme while another, encoded by an adjacent gene, is a short peptide predicted to be its substrate for posttranslational modification. The most prevalent, in over 125 genomes, featuring a peptide that we designate SCIFF (six cysteines in forty-five residues), is conserved throughout the class Clostridia, a distribution inconsistent with putative bacteriocin activity. A second novel system features a tandem pair of putative peptide-modifying radical SAM enzymes associated with a highly divergent family of peptides in which the only clearly conserved feature is a run of His-Xaa-Ser repeats. A third system pairs a radical SAM domain peptide maturase with selenocysteine-containing targets, suggesting a new biological role for selenium. These and several additional novel maturases that cooccur with predicted target peptides share a C-terminal additional 4Fe4S-binding domain with PqqE, the subtilosin A maturase AlbA, and the predicted mycofactocin and Nif11-class peptide maturases as well as with activators of anaerobic sulfatases and quinohemoprotein amine dehydrogenases. Radical SAM enzymes with this additional domain, as detected by TIGR04085, significantly outnumber lantibiotic synthases and cyclodehydratases combined in reference genomes while being highly enriched for members whose apparent targets are small peptides. Interpretation of comparative genomics evidence suggests unexpected (nonbacteriocin) roles for natural products from several of these systems.

Expansion of ribosomally produced natural products: a nitrile hydratase- and Nif11-related precursor family.

BACKGROUND: A new family of natural products has been described in which cysteine, serine and threonine from ribosomally-produced peptides are converted to thiazoles, oxazoles and methyloxazoles, respectively. These metabolites and their biosynthetic gene clusters are now referred to as thiazole/oxazole-modified microcins (TOMM). As exemplified by microcin B17 and streptolysin S, TOMM precursors contain an N-terminal leader sequence and C-terminal core peptide. The leader sequence contains binding sites for the posttranslational modifying enzymes which subsequently act upon the core peptide. TOMM peptides are small and highly variable, frequently missed by gene-finders and occasionally situated far from the thiazole/oxazole forming genes. Thus, locating a substrate for a particular TOMM pathway can be a challenging endeavor. RESULTS: Examination of candidate TOMM precursors has revealed a subclass with an uncharacteristically long leader sequence closely related to the enzyme nitrile hydratase. Members of this nitrile hydratase leader peptide (NHLP) family lack the metal-binding residues required for catalysis. Instead, NHLP sequences display the classic Gly-Gly cleavage motif and have C-terminal regions rich in heterocyclizable residues. The NHLP family exhibits a correlated species distribution and local clustering with an ABC transport system. This study also provides evidence that a separate family, annotated as Nif11 nitrogen-fixing proteins, can serve as natural product precursors (N11P), but not always of the TOMM variety. Indeed, a number of cyanobacterial genomes show extensive N11P paralogous expansion, such as Nostoc, Prochlorococcus and Cyanothece, which replace the TOMM cluster with lanthionine biosynthetic machinery. CONCLUSIONS: This study has united numerous TOMM gene clusters with their cognate substrates. These results suggest that two large protein families, the nitrile hydratases and Nif11, have been retailored for secondary metabolism. Precursors for TOMMs and lanthionine-containing peptides derived from larger proteins to which other functions are attributed, may be widespread. The functions of these natural products have yet to be elucidated, but it is probable that some will display valuable industrial or medical activities.

Analysis of rare genomic changes does not support the unikont-bikont phylogeny and suggests cyanobacterial symbiosis as the point of primary radiation of eukaryotes.

The deep phylogeny of eukaryotes is an important but extremely difficult problem of evolutionary biology. Five eukaryotic supergroups are relatively well established but the relationship between these supergroups remains elusive, and their divergence seems to best fit a "Big Bang" model. Attempts were made to root the tree of eukaryotes by using potential derived shared characters such as unique fusions of conserved genes. One popular model of eukaryotic evolution that emerged from this type of analysis is the unikont-bikont phylogeny: The unikont branch consists of Metazoa, Choanozoa, Fungi, and Amoebozoa, whereas bikonts include the rest of eukaryotes, namely, Plantae (green plants, Chlorophyta, and Rhodophyta), Chromalveolata, excavates, and Rhizaria. We reexamine the relationships between the eukaryotic supergroups using a genome-wide analysis of rare genomic changes (RGCs) associated with multiple, conserved amino acids (RGC_CAMs and RGC_CAs), to resolve trifurcations of major eukaryotic lineages. The results do not support the basal position of Chromalveolata with respect to Plantae and unikonts or the monophyly of the bikont group and appear to be best compatible with the monophyly of unikonts and Chromalveolata. Chromalveolata show a distinct, additional signal of affinity with Plantae, conceivably, owing to genes transferred from the secondary, red algal symbiont. Excavates are derived forms, with extremely long branches that complicate phylogenetic inference; nevertheless, the RGC analysis suggests that they are significantly more likely to cluster with the unikont-Chromalveolata assemblage than with the Plantae. Thus, the first split in eukaryotic evolution might lie between photosynthetic and nonphotosynthetic forms and so could have been triggered by the endosymbiosis between an ancestral unicellular eukaryote and a cyanobacterium that gave rise to the chloroplast.