DNA-bridging by an archaeal histone variant via a unique tetramerisation interface

In eukaryotes, histone paralogues form obligate heterodimers such as H3/H4 and H2A/H2B that assemble into octameric nucleosome particles. Archaeal histones are dimeric and assemble on DNA into 'hypernucleosome' particles of varying sizes with each dimer wrapping 30 bp of DNA. These are composed of canonical and variant histone paralogues, but the function of these variants is poorly understood. Here, we characterise the structure and function of the histone paralogue MJ1647 from Methanocaldococcus jannaschii that has a unique C-terminal extension enabling homotetramerisation. The 1.9 Å X-ray structure of a dimeric MJ1647 species, structural modelling of the tetramer, and site-directed mutagenesis reveal that the C-terminal tetramerization module consists of two alpha helices in a handshake arrangement. Unlike canonical histones, MJ1647 tetramers can bridge two DNA molecules in vitro. Using single-molecule tethered particle motion and DNA binding assays, we show that MJ1647 tetramers bind ~60 bp DNA and compact DNA in a highly cooperative manner. We furthermore show that MJ1647 effectively competes with the transcription machinery to block access to the promoter in vitro. To the best of our knowledge, MJ1647 is the first histone shown to have DNA bridging properties, which has important implications for genome structure and gene expression in archaea

Blackcaps fruit consumption

fruit consumption among blackcaps in 5 water treatments: 1. ad libitum MAMT fruits, but only for five hours each day + saline solution of 0.6 ml NaCl 0.9% subcutaneous injection. 2. simultaneous provisioning of fruits and water for 5 hours. 3. A group provided with drinking water for five hours, and after a one-hour break, provided with MAMT fruits for five hours 4 .A group provided with MAMT fruits for five hours, and after a one-hour break, provided with drinking water for five hours. 5. A water restricted group that was provided with MAMT fruits for five hours (n = 2)

68Ga-PSMA-11 PET/CT Follow-Up of Patients with Prostate Cancer with Bone Metastases Who Had Reduced Bone Density after Androgen Deprivation Therapy

Bone metastases from prostate cancer (PCa) often show an increase in density on computed tomography (CT) after successful androgen deprivation therapy (ADT). Density may be reduced, however, as the disease progresses or, contrarily, when disease is no longer active. The current study investigated the role of 68Ga-PSMA-11 positron emission tomography/computed tomography (PET/CT) in differentiating between these two conditions. Methods: The study cohort included 15 PCa patients with sclerotic/blastic bone metastasis in whom reduction in bone density of metastasis was noted on follow-up 68Ga-PSMA-11 PET/CT after ADT. Each patient had two PET/CT scans. Prior to the first scan, six patients were castration naïve and nine patients were already treated. All patients had ADT between the two PET/CT scans. PET parameters (SUVmax and tumor-to-background ratio), and CT parameters (HUmax) were determined and compared for each lesion on both scans. Patient’s response was based on prostate-specific antigen (PSA) levels and appearance of new lesions. The Kolmogorov–Smirnov test was used to evaluate normal distribution of the continuous variables. Results: Post-ADT reduction in bone density was identified in 37 lesions. The mean HUmax was 883.9 ± 175.1 on the first scan and 395.6 ± 157.1 on the second scan (p < 0.001). Twenty-one of the 37 lesions showed no increased tracer uptake on the second PET/CT scan raising the likelihood of a response. The other 16 lesions were associated with increased uptake suggestive of an active resistant disease. Bone density was not different in lesions that no longer showed an increased uptake as compared with those that did. Seven of the study patients responded to therapy, and none of the 16 lesions found in these patients showed increased 68Ga-PSMA-11 uptake. In eight patients with progressive disease, all 12 lesions in five of them showed increased 68Ga-PSMA-11 uptake, there was mixed response in two patients (having two lesions with increased uptake and one without) and although all three lesions no longer showed an increased uptake, new lesions were detected in the eighth patient. Conclusion: A decrease in density of bone lesions may reflect clinical progression, or contrarily, a response to therapy in patients with PCa and skeletal involvement treated with ADT. Uptake of 68Ga-PSMA-11 may separate between these two vastly opposing conditions

Guide-independent DNA cleavage by archaeal Argonaute from Methanocaldococcus jannaschii

Prokaryotic Argonaute proteins acquire guide strands derived from invading or mobile genetic elements, via an unknown pathway, to direct guide-dependent cleavage of foreign DNA. Here, we report that Argonaute from the archaeal organism Methanocaldococcus jannaschii (MjAgo) possesses two modes of action: the canonical guide-dependent endonuclease activity and a non-guided DNA endonuclease activity. The latter allows MjAgo to process long double-stranded DNAs, including circular plasmid DNAs and genomic DNAs. Degradation of substrates in a guide-independent fashion primes MjAgo for subsequent rounds of DNA cleavage. Chromatinized genomic DNA is resistant to MjAgo degradation, and recombinant histones protect DNA from cleavage in vitro. Mutational analysis shows that key residues important for guide-dependent target processing are also involved in guide-independent MjAgo function. This is the first characterization of guide-independent cleavage activity for an Argonaute protein potentially serving as a guide biogenesis pathway in a prokaryotic system

Excavations at Motza in the Judean Hills and the Early Pre-Pottery Neolithic B in the Southern Levant

International audienceRecent excavations at Motza near Jerusalem revealed a large Neolithic site that was continuously inhabited from the EarlyPPNB until the Pottery Neolithic period. The well-dated Early PPNB occupation comprises various architectural remains, human burials, clay and stone figurines, and rich flint and obsidian assemblages. Layer VI is approximately 2 m thick and consists of three discrete occupational levels with distinct architectural remains. These include curvilinear and rectangular structures with lime plastered floors, courtyards, hearths, and long massive walls of up to 1 m width. This layer is dated by sixteen radiocarbon dates, all falling within the range of 8600-8200 cal. BC. The EPPNB flint assemblage at Motza demonstrates a combination of the continuation of the earlier PPNA traditions together with the introduction of new technological innovations. The formal tools are dominated by Helwan and Jericho points, with many of the former being made on bidirectional blades in the north Levantine mode. Sickle blades are also mainly fashioned on bidirectional blanks. Mountain gazelle (Gazella gazella), the dominant species of the EPPNB faunal remains, indicates that hunting was still important. Gazelle bones were preferred for manufacturing bone tools, continuing PPNA Sultanian practices. Most of the human remains were primary burials, with no preference in the orientation of the bodies, although they were tightly flexed. Graves illustrate various funerary practices although primary inhumations on tightly flexed position are more common. All three primary graves of adults show evidence of later skull removal. Corpse treatment seems nonetheless quite particular at Motza as most of the graves were associated in different ways with remains of plaster, more frequently, as patches mixed in the fi lling of the grave. The archaeological evidence from Motza revises the hypothesis held by some that the PPNB in this region only started in the MPPNB. In addition, the advanced naviform technology and Helwan points present at Motza appear to be chronological markers of this period and together with radiocarbon dates confi rm the existence of this stage of the PPNB in the southern Levant.Les fouilles effectuées récemment à Motza, près de Jérusalem, ont révélé la présence d’un important site néolithique occupé de manière continue du PPNB ancien au Néolithique céramique. Bien datée, l’occupation du PPNB ancien a livré de nombreux vestiges architecturaux, des sépultures, des fi gurines d’argile et de pierre, ainsi qu’une abondante industrie sur silex et obsidienne. Le niveau 6, épais de près de deux mètres, comporte trois couches d’occupation successives, présentant des restes architecturaux distincts. Ceux-ci incluent des structures curvilinéaires et rectangulaires avec des sols de chaux, des cours, des foyers et des murs massifs de plus d’un mètre d’épaisseur. Cette couche est datée par seize datations radiocarbone toutes comprises entre 8600 et 8200 av. J.-C. (cal.). L’industrie lithique de Motza présente une combinaison de traditions plus anciennes du PPNA associées à des innovations technologiques. Les pointes d’Helwan et de Jéricho dominent l’outillage et beaucoup sont façonnées sur des lames bidirectionnelles comme au Levant Nord. La plupart des lames de faucilles sont elles aussi façonnées sur des produits bipolaires. La gazelle (Gazella gazella) qui dominel’assemblage faunique du PPNB ancien indique que la chasse reste importante à cette période. La gazelle sert préférentiellement de support pour l’industrie osseuse, un choix hérité du Sultanien. Les pratiques funéraires sont variées, mais les inhumations sont majoritairement primaires et la position sur le côté en fl exion forcée domine. Dans les trois inhumations primaires d’adultes, les crânes ont été prélevés après décomposition. Le traitement des corps observé à Motza se singularise par la présence de plâtre associé aux sépultures de diverses manières, particulièrement des morceaux mêlés au remplissage. Les découvertes faites à Motza invitent à revoir l’hypothèse selon laquelle, dans cette région, le PPNB n’aurait débuté qu’avec sa phase moyenne (MPPNB). De plus, la technologie naviforme évoluée et les pointes d’Helwan présentes à Motza, apparaissent comme des marqueurs chronologiques de cette période ; associées aux datations C14, elles confi rment l’existence de cette phase ancienne du PPNB au Levant Sud

DNA-bridging by an archaeal histone variant via a unique tetramerisation interface

Abstract In eukaryotes, histone paralogues form obligate heterodimers such as H3/H4 and H2A/H2B that assemble into octameric nucleosome particles. Archaeal histones are dimeric and assemble on DNA into ‘hypernucleosome’ particles of varying sizes with each dimer wrapping 30 bp of DNA. These are composed of canonical and variant histone paralogues, but the function of these variants is poorly understood. Here, we characterise the structure and function of the histone paralogue MJ1647 from Methanocaldococcus jannaschii that has a unique C-terminal extension enabling homotetramerisation. The 1.9 Å X-ray structure of a dimeric MJ1647 species, structural modelling of the tetramer, and site-directed mutagenesis reveal that the C-terminal tetramerization module consists of two alpha helices in a handshake arrangement. Unlike canonical histones, MJ1647 tetramers can bridge two DNA molecules in vitro. Using single-molecule tethered particle motion and DNA binding assays, we show that MJ1647 tetramers bind ~60 bp DNA and compact DNA in a highly cooperative manner. We furthermore show that MJ1647 effectively competes with the transcription machinery to block access to the promoter in vitro. To the best of our knowledge, MJ1647 is the first histone shown to have DNA bridging properties, which has important implications for genome structure and gene expression in archaea