Linker histones in hormonal gene regulation.

In the present review, we summarize advances in our knowledge on the role of the histone H1 family of proteins in breast cancer cells, focusing on their response to progestins. Histone H1 plays a dual role in gene regulation by hormones, both as a structural component of chromatin and as a dynamic modulator of transcription. It contributes to hormonal regulation of the MMTV promoter by stabilizing a homogeneous nucleosome positioning, which reduces basal transcription whereas at the same time promoting progesterone receptor binding and nucleosome remodeling. These combined effects enhance hormone dependent gene transcription, which eventually requires H1 phosphorylation and displacement. Various isoforms of histone H1 have specific functions in differentiated breast cancer cells and compact nucleosomal arrays to different extents in vitro. Genome-wide studies show that histone H1 has a key role in chromatin dynamics of hormone regulated genes. A complex sequence of enzymatic events, including phosphorylation by CDK2, PARylation by PARP1 and the ATP-dependent activity of NURF, are required for H1 displacement and gene de-repression, as a prerequisite for further nucleosome remodeling. Similarly, during hormone-dependent gene repression a dedicated enzymatic mechanism controls H1 deposition at promoters by a complex containing HP1γ, LSD1 and BRG1, the ATPase of the BAF complex. Thus, a broader vision of the histone code should include histone H1, as the linker histone variants actively participate in the regulation of the chromatin structure. How modifications of the core histones tails affect H1 modifications and vice versa is one of the many questions that remains to be addressed to provide a more comprehensive view of the histone cross-talk mechanisms.

Hypertrophic cardiomyopathy in a dog: a systematic diagnostic approach.

A seven-year-old female neutered Parson Russel terrier was referred for syncopal episodes. An electrocardiogram revealed paroxysmal atrial flutter followed by periods of sinus arrest, suggesting sick sinus syndrome. Echocardiography showed severe biventricular wall thickening (hypertrophic cardiomyopathy (HCM) phenotype) with no signs of fixed or dynamic left ventricular outflow tract obstruction. Blood pressure, abdominal ultrasound, serum total thyroxin and thyroid-stimulating hormone, and insulin-like growth factor-1 were all within normal limits. Cardiac troponin I was elevated (1.7 ng/mL, ref<0.07). Serological tests for common infectious diseases were negative. A 24-h Holter confirmed that the syncopal episodes were associated with asystolic pauses (sinus arrest after runs of atrial flutter) ranging between 8.5 and 9.6 s. Right ventricular endomyocardial biopsies (EMB) were performed at the time of pacemaker implantation to assess for storage or infiltrative diseases that mimic HCM in people. Histological analysis of the EMB revealed plurifocal inflammatory infiltrates with macrophages and lymphocytes (CD3+ > 7/mm2) associated with myocyte necrosis, but no evidence of myocyte vacuolisation or infiltrative myocardial disorders. These findings were compatible with myocardial ischaemic injury or acute lymphocytic myocarditis. Molecular analysis of canine cardiotropic viruses were negative. The dog developed refractory congestive heart failure and was euthanised 16 months later. Cardiac post-mortem examination revealed cardiomyocyte hypertrophy and disarray with diffuse interstitial and patchy replacement fibrosis, and small vessel disease, confirming HCM. We described a systemic diagnostic approach to an HCM phenotype in a dog, where a diagnosis of HCM was reached by excluding HCM phenocopies.

Reliable evaluation of the quantal determinants of synaptic efficacy using Bayesian analysis.

Communication between neurones in the central nervous system depends on synaptic transmission. The efficacy of synapses is determined by pre- and postsynaptic factors that can be characterized using quantal parameters such as the probability of neurotransmitter release, number of release sites, and quantal size. Existing methods of estimating the quantal parameters based on multiple probability fluctuation analysis (MPFA) are limited by their requirement for long recordings to acquire substantial data sets. We therefore devised an algorithm, termed Bayesian Quantal Analysis (BQA), that can yield accurate estimates of the quantal parameters from data sets of as small a size as 60 observations for each of only 2 conditions of release probability. Computer simulations are used to compare its performance in accuracy with that of MPFA, while varying the number of observations and the simulated range in release probability. We challenge BQA with realistic complexities characteristic of complex synapses, such as increases in the intra- or intersite variances, and heterogeneity in release probabilities. Finally, we validate the method using experimental data obtained from electrophysiological recordings to show that the effect of an antagonist on postsynaptic receptors is correctly characterized by BQA by a specific reduction in the estimates of quantal size. Since BQA routinely yields reliable estimates of the quantal parameters from small data sets, it is ideally suited to identify the locus of synaptic plasticity for experiments in which repeated manipulations of the recording environment are unfeasible.

Antigenic and genetic analyses of isolate APMV/wigeon/Italy/3920-1/2005 indicate that it represents a new avian paramyxovirus (APMV-12).

Isolate wigeon/Italy/3920-1/2005 (3920-1) was obtained during surveillance of wild birds in November 2005 in the Rovigo province of Northern Italy and shown to be a paramyxovirus. Analysis of cross-haemagglutination-inhibition tests between 3920-1 and representative avian paramyxoviruses showed only a low-level relationship to APMV-1. Phylogenetic analysis of the whole genome and each of the six genes indicated that while 3920-1 grouped with APMV-1 and APMV-9 viruses, it was quite distinct from these two. In the whole-genome analysis, 3920-1 had 52.1 % nucleotide sequence identity to the closest APMV-1 virus, 50.1 % identity to the APMV-9 genome, and less than 42 % identity to representatives of the other avian paramyxovirus groups. We propose isolate wigeon/Italy/3920-1/2005 as the prototype strain of a further APMV group, APMV-12.

An SYBR Green-based real-time RT-PCR assay for the detection of H5 hemagglutinin subtype avian influenza virus.

Increasing diversity among H5 hemagglutinin (HA) subtype avian influenza (AI) viruses has resulted in the need of novel sensitive and specific molecular assays. In this study, an SYBR Green-based real-time reverse transcription-PCR (RRT-PCR) assay was developed for the detection of H5 subtype AI virus. Sequence analysis of the Mexican lineage H5N2 isolates (subgroup B) revealed several mismatches in the primer/hydrolysis probe set reported in the commonly used RRT-PCR assay for the detection of H5 North American lineage. The present assay was designed to circumvent the challenge that these viruses represent for the specific detection of H5 subtype AI viruses. This RRT-PCR assay successfully detected a range of different H5 subtype AI strains from both Eurasian and North American lineages representing different avian H5 HA clades from diverse geographical locations. The sensitivity of the present method was determined by using in vitro-transcribed RNA and 10-fold serial dilutions of titrated AI viruses. High sensitivity levels were obtained, with limits of detection of 10(0) 50% egg infectious dose (EID50)/mL and 4.2 gene copies/µl. The linear ranges of the assay span within 10(6)-10(0) EID50/mL and 10(6)-10(0) gene copies/µl. The results obtained from this method were directly compared with those of the H5 RRT-PCR assay recommended by the OIE. The comparison was performed with 110 tracheal and cloacal swabs from various bird species collected during field and laboratory investigations in Eurasia and Africa in 2006 and 2008 and showed 100% agreement. This assay is recommended as an alternative method, also allowing a 'double check' approach detection, to be use mainly in outbreak scenarios with higher risk of poultry infections by Central American/Caribbean H5 AI viruses.

Nodular lesions arising in a large congenital melanocytic naevus in a newborn with eruptive disseminated Spitz naevi.

Congenital malignant melanoma within a pre-existing large congenital melanocytic naevus (CMN) is exceedingly rare. Its incidence is difficult to determine due to the small number of reported cases and because of problems associated with diagnosis. Some benign nodular proliferations (called proliferative nodules) arising in CMN, while rare, are significantly more common and can mimic malignant melanoma clinically or histologically. There are no reported cases of congenital melanoma or benign proliferative nodules in CMN in patients who also had eruptive disseminated Spitz naevi. We describe a girl who was noted to have a dark-brown plaque with several large erythematous nodules affecting the scalp at delivery, in addition to multiple erythematous dome-shaped papules that developed in a disseminated manner over several months, beginning at 10 days of age. It was difficult, not only clinically but also histologically, to determine the benign or malignant nature of all of these lesions. As primary cutaneous melanoma, atypical proliferative nodules in CMN, bland CMN or CMN with foci of increased cellularity and Spitz naevi show clear differences in the genetic aberration patterns, comparative genomic hybridization (CGH) could be a diagnostic help in ambiguous cases such as this. CGH performed on this patient showed multiple DNA copy number changes in the most atypical nodule, but such alterations could not be found in the remainder of the lesions. CGH showed differences between the nodular lesions that occurred in the CMN and helped us in supporting the diagnosis of this unique case of benign proliferative nodules and a possible congenital melanoma arising in a large CMN, associated with multiple widespread eruptive Spitz naevi.

Transboundary spread of highly pathogenic avian influenza through poultry commodities and wild birds: a review.

The extensive circulation of the H5N1 highly pathogenic avian influenza (HPAI) virus in animals and the human health implications which it poses have led to extensive research in unexplored fields and thus a re-assessment of our understanding of this infection. Moreover, widespread infection of poultry has raised concerns about the food safety and trade implications of this infection, necessitating revised international trade regulations. The role of wild birds has been much debated and resources have been invested to clarify the role that they may play in the spread of infection. It is now clear that wild birds may be responsible for primary introduction in a previously free area. To date it is still unclear whether HPAI infection may be maintained in wild bird populations for extended periods of time. This paper reviews existing knowledge on the transboundary spread of HPAI through poultry and poultry commodities and summarises evidence of spread through wild birds.

Multifocal lymphangioendotheliomatosis with thrombocytopenia: phenotypic variability and response to rapamycin. / Linfagioendoteliomatosis multifocal con trombopenia: variabilidad fenotípica y respuesta a rapamicina.

INTRODUCTION: Multifocal lymphangioendotheliomatosis with thrombocytopenia (MLT) is characterized by multiple maculopapular lesions involving the stomach and the lungs, associated with thrombocytopenia as a result of platelet entrapment. Episodes of severe digestive bleeding, which are sometimes unmanageable, are one of its most frequent presentations and a cause of mortality. Our objective was to describe the various phenotypes, as well as our treatment experience. MATERIALS AND METHODS: A retrospective analysis of patients diagnosed with MLT in our vascular abnormality unit from 2007 to 2018 was carried out. Epidemiological, clinical, and evolution data were analyzed, and a long-term follow-up was performed. RESULTS: Five patients (3 boys and 2 girls) had congenital macules and erythematous papules of various sizes. They were later associated with episodes of severe hematemesis along with thrombocytopenia, which required blood product transfusion. The most frequently involved areas were the stomach and the colon. In two patients, multiple bilateral pulmonary nodules were noted. The anatomical pathology examination showed extended vessels with a prominent, hobnail endothelium, as well as intraluminal papillary projections in the dermis. Immunohistochemical analysis was CD-31 positive and CD-34 positive in a characteristic manner. Two patients were treated with mTOR inhibitors (rapamycin), with a progressive decrease in extracutaneous involvement and platelet recovery, but with a poor response in dermal lesions. Two patients were treated with vincristine, with a reduction of digestive bleeding episodes. No deaths were reported in our series. CONCLUSION: MLT is characterized by hematological and cutaneous involvement - sometimes minimal -, with potential lesions in other internal organs. Its heterogeneous presentation, which may start with severe digestive bleeding, makes this rare pathology difficult to diagnose. mTOR inhibitors have opened up new treatment possibilities.

INTRODUCCION: La linfangioendoteliomatosis multifocal con trombopenia (LMT) es una anomalía, caracterizada por múltiples lesiones maculo-papulosas con afectación visceral gástrica y pulmonar, asociado a trombopenia por atrapamiento plaquetar. Una de sus presentaciones más frecuentes es en forma de episodios de hemorragia digestiva severa, en ocasiones inmanejable, y que es la responsable de su mortalidad. Nuestro objetivo es describir los diferentes fenotipos, así como nuestra experiencia en su tratamiento. MATERIAL Y METODOS: Hemos realizado un análisis retrospectivo de los pacientes diagnósticos de LMT según las características histológicas típicas entre 2007 y 2018 en nuestra unidad de anomalías vasculares. Se analizaron datos epidemiológicos, clínicos y de evolución, así como seguimiento a largo plazo. RESULTADOS: Cinco pacientes (3 hombres y 2 mujeres) presentaron al nacimiento máculas y pápulas eritematosas de diferentes tamaños a los que más adelante se les asoció episodios de hematemesis graves junto a trombopenia, que llegaron a requerir transfusión de hemoderivados. Las regiones más afectadas fueron el estómago seguido del colon. En dos pacientes se detectaron múltiples nódulos pulmonares bilaterales. La anatomía patológica describió vasos alargados con endotelio prominente y en tachuela junto a proyecciones papilares intraluminales en dermis. La inmunohistoquímica fue positiva de forma característica para CD-31 y CD-34. Dos pacientes fueron tratados con inhibidores de mTOR (rapamicina) con disminución progresiva de la afectación extracutánea y recuperación plaquetar, pero con una pobre respuesta de las lesiones dérmicas. Dos pacientes fueron tratados con vincristina con reducción de los episodios de sangrado digestivo. No se registró ningún fallecimiento en nuestra serie. CONCLUSION: La LMT se caracteriza por una afectación cutánea, a veces mínima, y hematológica que puede asociar lesiones en otros órganos internos. La presentación heterogénea, pudiendo debutar con hemorragias digestivas severas, hacen de esta entidad una patología de difícil diagnóstico. Los inhibidores de mTOR han abierto una nueva vía que arroja cierta esperanza para el tratamiento de esta patología tan poco frecuente.

Author Correction: Unrevealed genetic diversity of GII Norovirus in the swine population of North East Italy.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Unrevealed genetic diversity of GII Norovirus in the swine population of North East Italy.

Noroviruses (NoVs) are one of the major causative agents of non-bacterial gastroenteritis in humans worldwide. NoVs, belonging to Caliciviridae, are classified into ten genogroups (G) and eight P-groups based on major capsid protein (VP1) and of the RNA-dependent-RNA-polymerase (RdRp), respectively. In swine, the main genogroup and P-group identified are GII and GII.P; which can infect humans too. To date, only one case of GIIP.11 have been identified in swine in Italy while the circulation of other P-types is currently unknown. In the present study, 225 swine faecal samples were collected from 74 swine herds in Veneto region through on-farm monitoring. NoV circulation was particularly high in older pigs. The phylogenetic analysis showed the co-circulation of NoVs belonging to two different P-types: GII.P11 and GII.P18, here described for the first time in Italy, presenting an extensive genetic diversity, never described before worldwide. Distinct NoV genetic subgroups and unique amino acid mutations were identified for each P-type for the first time. This study demonstrated the co-circulation of diverse swine NoVs subgroups in Italy, raising questions on the origin of such diversity and suggesting that continuous monitoring of swine NoVs is needed to track the emergence of potentially zoonotic viruses by recombination events.

Negative regulation by glucocorticoids through interference with a cAMP responsive enhancer.

Although steroid hormone receptors are known to activate gene expression by binding to specific hormone-dependent enhancers, the mechanisms by which steroids inhibit the transcription of specific genes are unknown. It is shown here by gene transfer studies that the same glucocorticoid receptor that activates gene expression can negatively regulate expression of the human glycoprotein hormone alpha-subunit gene. Glucocorticoid inhibition was conferred by a 52-nucleotide region that also contains elements crucial both for adenosine 3',5'-monophosphate (cAMP) responsiveness and for placental-specific expression of this gene and was observed only under conditions in which these elements were functioning as enhancers. Purified glucocorticoid receptor was found to bind to DNA that overlap the cAMP responsive elements sites in this region. It is hypothesized that steroid receptors negatively regulate gene expression by interfering with the activity or binding of other important transcription factors.

Nearly Complete Genome Sequence of a Sapelovirus A Strain Identified in Swine in Italy.

We report the first nearly complete genome sequence of a porcine sapelovirus (PSV) A strain that was identified from feces of piglets suffering from diarrhea in Italy in 2015. Phylogenetic investigations revealed a separate clustering for the Italian PSV, indicating unique molecular features.

Epidemiology and natural history of cutaneous squamous cell carcinoma in recessive dystrophic epidermolysis bullosa patients: 20 years' experience of a reference centre in Spain.

BACKGROUND: Cutaneous squamous cell carcinoma (cSCC) is the leading cause of death in patients with recessive dystrophic epidermolysis bullosa (RDEB). We provide the management and prognosis of cSCC in RDEB patients at a Spanish reference center. MATERIALS AND METHODS: We retrospectively included patients with RDEB attended in La Paz University Hospital from November 1988 to October 2018. RESULTS: Fourteen patients developed at least one cSCC. Tumors were predominantly well differentiated. Nearly half of the tumors have recurred. Median time to first recurrence was 23.4 months (95% CI: 17.2-29.5). Five patients have developed distant metastases. Median overall survival (mOS) was 136.5 months since the diagnosis of the first cSCC (95% CI: 30.6-242.3). When distant metastases occurred, mOS was 6.78 months (95% CI: 1.94-11.61). CONCLUSIONS: cSCC is a life-threatening complication of RDEB patients. Although tumors are usually well differentiated, they tend to relapse. This is the first Spanish report of cSCC arising in RDEB patients.

Steroid hormone receptors: interaction with deoxyribonucleic acid and transcription factors.

Gene regulation by steroid hormones is mediated by binding of the hormone ligand to the corresponding receptor that triggers a complex set of interactions of the hormone receptors with each other, with DNA in chromatin, and with a variety of other proteins. In this review we attempt to summarize what is known about these interactions using as the main example the regulation of mouse mammary tumor virus transcription by glucocorticoids and progestins. We describe in some detail the interaction of monomers and homodimers of the steroid receptors with their recognition sequences, and the molecular mechanism used to discriminate between the responsive elements for glucocorticoids/progestins and estrogens. We then review the interactions between homologous and heterologous hormone receptors on complex hormone regulatory regions, before devoting some attention to the synergistic and inhibitory interactions of hormone receptors with other transcription factors. Finally we briefly summarize some of the possible mechanisms that modulate the molecular interactions of hormone receptors. In addition to ligand binding, these include receptor phosphorylation, changes in DNA topology, and the organization of DNA in nucleosomes. From this overview we draw the tentative conclusion that the specificity of the hormonal response in different cells results from a combination of developmental restrictions both in the accessibility of genomic sequence and in the repertoire of regulatory proteins present in each particular cell. In addition, the array of regulatory sequences in DNA and chromatin determines the precise nature of macromolecular interactions of the receptors that are modulated by their degree of phosphorylation.

Hodgkin's lymphoma cells express alternatively spliced forms of HDM2 with multiple effects on cell cycle control.

The HDM2 oncoprotein is a cellular inhibitor of p53 and is frequently deregulated in human cancer. However, the HDM2 gene encodes alternatively spliced variants whose functional significance is poorly understood. We had previously reported the detection of alternative HDM2 forms in Hodgkin's lymphoma (HL)-derived cell lines. Here, we have cloned several of these transcripts, including the previously described HDM2-A, -B and -C (which encode the COOH terminus of HDM2), and two novel variants (HDM2-HL1 and -HL2) containing a complete p53 interaction domain. Real-time PCR assays demonstrated that HDM2-A and -B were selectively expressed by HL cell lines and primary tumors, compared with their non-neoplastic counterparts. In transient transfection experiments, alternatively spliced HDM2 isoforms were partially or totally localized within the cytoplasm. HDM2-HL2 was able to inhibit transactivation of a p53-inducible reporter construct and induced a partial relocalization of p53 to the cytoplasm. Expression of HDM2-A and -B caused the activation of p53/p21 and induced growth arrest in primary cells, but also increased the expression levels of cyclins D1 and E. Other possible genes regulated by HDM2-A and -B were identified using cDNA microarray technology. These results imply that HDM2 isoforms may have multiple effects on cell cycle control, and provide insight into the mechanisms through which these molecules contribute to tumorigenesis.

Binding of YY1 to a site overlapping a weak TATA box is essential for transcription from the uteroglobin promoter in endometrial cells.

The gene for rabbit uteroglobin codes for a small calcium-, steroid-, and biphenyl metabolite-binding homodimeric protein which is expressed in a variety of epithelial cell types such as Clara cells (lung) and the glandular and luminal cells of the endometrium. One important region mediating its efficient transcription in a human endometrium-derived cell line, Ishikawa, is centered around a noncanonical TATA box. Two factors, TATA core factor (TCF), expressed in cell lines derived from uteroglobin-expressing tissues, and the ubiquitously expressed TATA palindrome factor, bind to the DNA major groove at two adjacent sites within this region. Here, we report the identification of the TATA palindrome factor as the transcription/initiation factor YY1 by microsequencing of the biochemically purified factor from HeLa cells. The binding site for YY1 within the uteroglobin gene is unique in its sequence and its location overlapping a weak TATA box (TACA). Binding of YY1 was required for efficient transcription in TCF-positive Ishikawa cells, which responded only weakly to a change of TACA to TATA, although in vitro binding affinity for the TATA-box-binding protein increased by 1 order of magnitude. In contrast, in CV-1 cells, lacking TCF, binding of YY1 was not required for transcription in the context of a wild-type TACA box, whereas a change from TACA to TATA led to significantly increased reporter gene expression. DNA binding data exclude a role of YY1 in stabilizing the interaction of the TATA-box-binding protein with the uteroglobin promoter. We conclude that cell lines derived from uteroglobin-expressing tissues overcome the weak TATA box with the help of auxiliary factors, one of them being YY1.