Vertebrate centromeres in mitosis are functionally bipartite structures stabilized by cohesin.

Centromeres are scaffolds for the assembly of kinetochores that ensure chromosome segregation during cell division. How vertebrate centromeres obtain a three-dimensional structure to accomplish their primary function is unclear. Using super-resolution imaging, capture-C, and polymer modeling, we show that vertebrate centromeres are partitioned by condensins into two subdomains during mitosis. The bipartite structure is found in human, mouse, and chicken cells and is therefore a fundamental feature of vertebrate centromeres. Super-resolution imaging and electron tomography reveal that bipartite centromeres assemble bipartite kinetochores, with each subdomain binding a distinct microtubule bundle. Cohesin links the centromere subdomains, limiting their separation in response to spindle forces and avoiding merotelic kinetochore-spindle attachments. Lagging chromosomes during cancer cell divisions frequently have merotelic attachments in which the centromere subdomains are separated and bioriented. Our work reveals a fundamental aspect of vertebrate centromere biology with implications for understanding the mechanisms that guarantee faithful chromosome segregation.

Successful anti-tumor effects with two novel bifunctional chemotherapeutic compounds that combine a LAT1 substrate with cytotoxic moieties in aggressive T-cell lymphomas.

T-cell lymphomas are aggressive neoplasms characterized by poor responses to current chemotherapeutic agents. Expression of the l-type amino acid transporter 1 (LAT 1, SLC7A5) allows for the expansion of healthy T-cell counterparts, and upregulation of LAT1 has been reported in precursor T-cell acute leukemia. Therefore, the expression of LAT1 was evaluated in a cohort of cutaneous and peripheral T-cell lymphomas. The findings demonstrated that LAT1 is upregulated in aggressive variants and absent in low-grade or indolent disease such as mycosis fungoides. In addition, upregulated LAT1 expression was seen in a large proportion of aggressive peripheral T-cell lymphomas, including peripheral T-cell lymphoma not otherwise specific (PTCL-NOS) and angioimmunoblastic T-cell lymphoma (AITL). The anti-tumor effects of two novel non-cleavable and bifunctional compounds, QBS10072S and QBS10096S, that combine a potent cytotoxic chemotherapeutic domain (tertiary N-bis(2-chloroethyl)amine) with the structural features of a selective LAT1 substrate (aromatic ß-amino acid) were tested in vitro and in vivo in T-cell lymphoma cell lines. The findings demonstrated decreased survival of T-cell lymphoma lines with both compounds. Overall, the results demonstrate that LAT1 is a valuable biomarker for aggressive T-cell lymphoma counterparts and QBS10072S and QBS10096S are successful therapeutic options for these aggressive diseases.

CD30 + Primary intestinal T-cell lymphoma (unclassified) masquerading as chronic inflammation: a case report.

BACKGROUND: Primary intestinal T-cell lymphomas are uncommon malignancies that pose a diagnostic dilemma, because the clinical features and imaging findings commonly overlap with those encountered in inflammatory bowel diseases. CASE PRESENTATION: The current clinical case report describes the clinical history, laboratory findings and histopathological analysis from a patient with non-specific gastrointestinal symptoms with a presumptive clinical diagnosis of inflammatory bowel disease, and two intestinal biopsy specimens with non-specific findings. Due to the persistent symptoms a third biopsy was consistent with primary intestinal T-cell lymphoma, a diagnosis that was elusive for months after the initial presentation. Clinical correlation with laboratory and histopathological findings is required to establish a definitive diagnosis and to further stratify the patients. In addition, the neoplastic cells featured partial expression of CD30, which had relevant therapeutic implications. CONCLUSIONS: Suspicion for an intestinal T-cell lymphoproliferative disorder should always exist in patients with persistent abdominal symptoms with no clear etiology. The current discussion provides a summary and review of the key diagnostic histological features for the classification of primary intestinal T-cell lymphomas. In addition, the discussion describes how specific the histological findings are relevant for the clinical management decisions.

Unique presentation of recurrent subdural effusions and subsequent abdominal lymphadenopathy in a patient diagnosed with Kikuchi-Fujimoto disease.

Kikuchi Fujimoto Disease, originally discovered in 1972, is a rare lymphoproliferative disorder traditionally characterized by cervical lymphadenopathy, fevers, parotid gland enlargement, and several other nonspecific manifestations. Differentials include lymphoma, other viral diseases such as Epstein-Bar Virus, as well as other autoimmune conditions such as Systemic Lupus Erythematosus. Central nervous system involvement is exceptionally rare, with manifestations including meningitis as well as subdural effusions, as presented in this case. This review will summarize a case of a 24-year-old man with recurrent subdural effusions requiring intervention, subsequent relapse with abdominal lymphadenopathy, and possible IgG4 related disease. The background epidemiology, radiology, and potential pathophysiology will be reviewed.

Activity of Lymphostatin, A Lymphocyte Inhibitory Virulence Factor of Pathogenic Escherichia coli, is Dependent on a Cysteine Protease Motif.

Lymphostatin (LifA) is a 366 kDa protein expressed by attaching & effacing Escherichia coli. It plays an important role in intestinal colonisation and inhibits the mitogen- and antigen-stimulated proliferation of lymphocytes and the synthesis of proinflammatory cytokines. LifA exhibits N-terminal homology with the glycosyltransferase domain of large clostridial toxins (LCTs). A DTD motif within this region is required for lymphostatin activity and binding of the sugar donor uridine diphosphate N-acetylglucosamine. As with LCTs, LifA also contains a cysteine protease motif (C1480, H1581, D1596) that is widely conserved within the YopT-like superfamily of cysteine proteases. By analogy with LCTs, we hypothesised that the CHD motif may be required for intracellular processing of the protein to release the catalytic N-terminal domain after uptake and low pH-stimulated membrane insertion of LifA within endosomes. Here, we created and validated a C1480A substitution mutant in LifA from enteropathogenic E. coli strain E2348/69. The purified protein was structurally near-identical to the wild-type protein. In bovine T lymphocytes treated with wild-type LifA, a putative cleavage product of approximately 140 kDa was detected. Appearance of the putative cleavage product was inhibited in a concentration-dependent manner by bafilomycin A1 and chloroquine, which inhibit endosome acidification. The cleavage product was not observed in cells treated with the C1480A mutant of LifA. Lymphocyte inhibitory activity of the purified C1480A protein was significantly impaired. The data indicate that an intact cysteine protease motif is required for cleavage of lymphostatin and its activity against T cells.

Red blood cell exchange to avoid intubating a COVID-19 positive patient with sickle cell disease?

As the COVID-19 pandemic continues to claim lives across the globe, insufficient data exists regarding the optimal treatment. It is well known that patients 55 years of age or older and patients with certain chronic diseases are at higher risk of severe illness, including acute respiratory distress syndrome and death. A potentially fatal pulmonary complication of sickle cell disease, acute chest syndrome, can be precipitated by acute infections, including respiratory viruses. We report the case of a patient with sickle cell disease (HbSC) who developed COVID-19 pneumonia and acute chest syndrome who was treated with emergent red blood cell exchange in order to avoid endotracheal intubation.

Loss of Tet1-Associated 5-Hydroxymethylcytosine Is Concomitant with Aberrant Promoter Hypermethylation in Liver Cancer.

Aberrant hypermethylation of CpG islands (CGI) in human tumors occurs predominantly at repressed genes in the host tissue, but the preceding events driving this phenomenon are poorly understood. In this study, we temporally tracked epigenetic and transcriptomic perturbations that occur in a mouse model of liver carcinogenesis. Hypermethylated CGI events in the model were predicted by enrichment of the DNA modification 5-hydroxymethylcytosine (5hmC) and the histone H3 modification H3K27me3 at silenced promoters in the host tissue. During cancer progression, selected CGIs underwent hypo-hydroxymethylation prior to hypermethylation, while retaining H3K27me3. In livers from mice deficient in Tet1, a tumor suppressor involved in cytosine demethylation, we observed a similar loss of promoter core 5hmC, suggesting that reduced Tet1 activity at CGI may contribute to epigenetic dysregulation during hepatocarcinogenesis. Consistent with this possibility, mouse liver tumors exhibited reduced Tet1 protein levels. Similar to humans, DNA methylation changes at CGI in mice did not appear to be direct drivers of hepatocellular carcinoma progression, rather, dynamic changes in H3K27me3 promoter deposition correlated strongly with tumor-specific activation and repression of transcription. Overall, our results suggest that loss of promoter-associated 5hmC in liver tumors licenses reprograming of DNA methylation at silent CGI during progression. Cancer Res; 76(10); 3097-108. ©2016 AACR.

A splicing-dependent transcriptional checkpoint associated with prespliceosome formation.

There is good evidence for functional interactions between splicing and transcription in eukaryotes, but how and why these processes are coupled remain unknown. Prp5 protein (Prp5p) is an RNA-stimulated adenosine triphosphatase (ATPase) required for prespliceosome formation in yeast. We demonstrate through in vivo RNA labeling that, in addition to a splicing defect, the prp5-1 mutation causes a defect in the transcription of intron-containing genes. We present chromatin immunoprecipitation evidence for a transcriptional elongation defect in which RNA polymerase that is phosphorylated at Ser5 of the largest subunit's heptad repeat accumulates over introns and that this defect requires Cus2 protein. A similar accumulation of polymerase was observed when prespliceosome formation was blocked by a mutation in U2 snRNA. These results indicate the existence of a transcriptional elongation checkpoint that is associated with prespliceosome formation during cotranscriptional spliceosome assembly. We propose a role for Cus2p as a potential checkpoint factor in transcription.

Cell type-specific DNA methylation at intragenic CpG islands in the immune system.

Human and mouse genomes contain a similar number of CpG islands (CGIs), which are discrete CpG-rich DNA sequences associated with transcription start sites. In both species, ∼50% of all CGIs are remote from annotated promoters but, nevertheless, often have promoter-like features. To determine the role of CGI methylation in cell differentiation, we analyzed DNA methylation at a comprehensive CGI set in cells of the mouse hematopoietic lineage. Using a method that potentially detects ∼33% of genomic CpGs in the methylated state, we found that large differences in gene expression were accompanied by surprisingly few DNA methylation changes. There were, however, many DNA methylation differences between hematopoietic cells and a distantly related tissue, brain. Altered DNA methylation in the immune system occurred predominantly at CGIs within gene bodies, which have the properties of cell type-restricted promoters, but infrequently at annotated gene promoters or CGI flanking sequences (CGI "shores"). Unexpectedly, elevated intragenic CGI methylation correlated with silencing of the associated gene. Differentially methylated intragenic CGIs tended to lack H3K4me3 and associate with a transcriptionally repressive environment regardless of methylation state. Our results indicate that DNA methylation changes play a relatively minor role in the late stages of differentiation and suggest that intragenic CGIs represent regulatory sites of differential gene expression during the early stages of lineage specification.

Orphan CpG islands identify numerous conserved promoters in the mammalian genome.

CpG islands (CGIs) are vertebrate genomic landmarks that encompass the promoters of most genes and often lack DNA methylation. Querying their apparent importance, the number of CGIs is reported to vary widely in different species and many do not co-localise with annotated promoters. We set out to quantify the number of CGIs in mouse and human genomes using CXXC Affinity Purification plus deep sequencing (CAP-seq). We also asked whether CGIs not associated with annotated transcripts share properties with those at known promoters. We found that, contrary to previous estimates, CGI abundance in humans and mice is very similar and many are at conserved locations relative to genes. In each species CpG density correlates positively with the degree of H3K4 trimethylation, supporting the hypothesis that these two properties are mechanistically interdependent. Approximately half of mammalian CGIs (>10,000) are "orphans" that are not associated with annotated promoters. Many orphan CGIs show evidence of transcriptional initiation and dynamic expression during development. Unlike CGIs at known promoters, orphan CGIs are frequently subject to DNA methylation during development, and this is accompanied by loss of their active promoter features. In colorectal tumors, however, orphan CGIs are not preferentially methylated, suggesting that cancer does not recapitulate a developmental program. Human and mouse genomes have similar numbers of CGIs, over half of which are remote from known promoters. Orphan CGIs nevertheless have the characteristics of functional promoters, though they are much more likely than promoter CGIs to become methylated during development and hence lose these properties. The data indicate that orphan CGIs correspond to previously undetected promoters whose transcriptional activity may play a functional role during development.