Trafficking of Estrella lausannensis in human macrophages.

Estrella lausannensis is a new member of the Chlamydiales order. Like other Chlamydia-related bacteria, it is able to replicate in amoebae and in fish cell lines. A preliminary study investigating the pathogenic potential of Chlamydia-related bacteria found a correlation between antibody response to E. lausannensis and pneumonia in children. To further investigate the pathogenic potential of E. lausannensis, we determined its ability to grow in human macrophages and its intracellular trafficking. The replication in macrophages resulted in viable E. lausannensis; however, it caused a significant cytopathic effect. The intracellular trafficking of E. lausannensis was analyzed by determining the interaction of the Estrella-containing inclusions with various endocytic markers as well as host organelles. The E. lausannensis inclusion escaped the endocytic pathway rapidly avoiding maturation into phagolysosomes by preventing both EEA-1 and LAMP-1 accumulation. Compared to Waddlia chondrophila, another Chlamydia-related bacteria, the recruitment of mitochondria and endoplasmic reticulum was minimal for E. lausannensis inclusions. Estrella lausannensis appears to use a distinct source of nutrients and energy compared to other members of the Chlamydiales order. In conclusion, we hypothesize that E. lausannensis has a restricted growth in human macrophages, due to its reduced capacity to control programmed cell death

Bilateral neglected posterior dislocation of the shoulder treated by reverse arthroplasty and contralateral osteochondral autograft. A case report

Bilateral posterior dislocation of the glenohumeral joint is an uncommon event, that can be missed at the initial presentation. We report the case of a 76-year old woman, who suffered a traumatic bilateral posterior dislocation, that was diagnosed three months later. She underwent surgical treatment on both shoulders in a single stage. Since the right shoulder showed a defect of the articular surface >50%, a reverse shoulder arthroplasty was performed on this side. The resected portion of the humeral head was retrieved and used as osteochondral graft to fill the reverse Hill-Sachs lesion of the left shoulder. At 18-month follow up, the patient was pain-free and had recovered excellent shoulder function on both sides: Constant score was 79 for the right shoulder and 88 for the left one. X-rays showed a grade 1 scapular notch of the right reverse prosthesis and good incorporation of the graft in the left shoulder, with no evidence of degenerative joint changes. Neglected posterior dislocations of the shoulder can be surgically treated by replacement or reconstruction. In case of bilateral injuries, the surgeon should carefully evaluate the pathoanatomy of both glenohumeral joints in order to choose and plan the most suitable procedure. If shoulder replacement is required on one side, the resected portion of the humeral head can be used as osteochondral autograft for a reconstruction procedure in the opposite side. The choice is influenced by several variables and decision-making might be challenging

Crescent and star shapes of members of the Chlamydiales order: impact of fixative methods.

Members of the Chlamydiales order all share a biphasic lifecycle alternating between small infectious particles, the elementary bodies (EBs) and larger intracellular forms able to replicate, the reticulate bodies. Whereas the classical Chlamydia usually harbours round-shaped EBs, some members of the Chlamydia-related families display crescent and star-shaped morphologies by electron microscopy. To determine the impact of fixative methods on the shape of the bacterial cells, different buffer and fixative combinations were tested on purified EBs of Criblamydia sequanensis, Estrella lausannensis, Parachlamydia acanthamoebae, and Waddlia chondrophila. A linear discriminant analysis was performed on particle metrics extracted from electron microscopy images to recognize crescent, round, star and intermediary forms. Depending on the buffer and fixatives used, a mixture of alternative shapes were observed in varying proportions with stars and crescents being more frequent in C. sequanensis and P. acanthamoebae, respectively. No tested buffer and chemical fixative preserved ideally the round shape of a majority of bacteria and other methods such as deep-freezing and cryofixation should be applied. Although crescent and star shapes could represent a fixation artifact, they certainly point towards a diverse composition and organization of membrane proteins or intracellular structures rather than being a distinct developmental stage

CRISPR System Acquisition and Evolution of an Obligate Intracellular Chlamydia-Related Bacterium.

Recently, a new Chlamydia-related organism, Protochlamydia naegleriophila KNic, was discovered within a Naegleria amoeba. To decipher the mechanisms at play in the modeling of genomes from the Protochlamydia genus, we sequenced the full genome of Pr. naegleriophila, which includes a 2,885,090 bp chromosome and a 145,285 bp megaplasmid. For the first time within the Chlamydiales order, we describe the presence of a clustered regularly interspaced short palindromic repeats (CRISPR) system, the immune system of bacteria, located on the chromosome. It is composed of a small CRISPR locus comprising eight repeats and associated cas-cse genes of the subtype I-E. A CRISPR locus is also present within Chlamydia sp. Diamant, another Pr. naegleriophila strain, suggesting that the CRISPR system was acquired by a common ancestor of Pr. naegleriophila, after its divergence from Pr. amoebophila. Both nucleotide bias and comparative genomics approaches identified probable horizontal gene acquisitions within two and four genomic islands in Pr. naegleriophila KNic and Diamant genomes, respectively. The plasmid encodes an F-type conjugative system highly similar to 1) that found in the Pam100G genomic island of Pr. amoebophila UWE25 chromosome, as well as on the plasmid of Rubidus massiliensis and 2) to the three genes remaining in the chromosome of Parachlamydia acanthamoebae strains. Therefore, this conjugative system was likely acquired on an ancestral plasmid before the divergence of Parachlamydiaceae Overall, this new complete Pr. naegleriophila genome sequence enables further investigation of the dynamic processes shaping the genomes of the family Parachlamydiaceae and the genus Protochlamydia

Functional consequences of mutations in CDKL5, an X-linked gene involved in infantile spasms and mental retardation

Mutations in the X-linked cyclin-dependent kinase-like 5 (CDKL5) gene have been identified in patients with Rett syndrome, West syndrome, and X-linked infantile spasms sharing the common features of generally intractable early seizures and mental retardation. Disease-causing mutations are distributed in both the catalytic domain and in the large COOH terminus. In this report, we examine the functional consequences of some Rett mutations of CDKL5 together with some synthetically designed derivatives useful to underline the functional domains of the protein. The mutated CDKL5 derivatives have been subjected to in vitro kinase assays and analyzed for phosphorylation of the TEY (Thr-Glu-Tyr) motif within the activation loop, their subcellular localization, and the capacity of CDKL5 to interact with itself. Whereas wild-type CDKL5 autophosphorylates and mediates the phosphorylation of the methyl-CpG-binding protein 2 (MeCP2) in vitro, Rett-mutated proteins show both impaired and increased catalytic activity suggesting that a tight regulation of CDKL5 is required for correct brain functions. Furthermore, we show that CDKL5 can self-associate and mediate the phosphorylation of its own TEY (Thr-Glu-Tyr) motif. Eventually, we show that the COOH terminus regulates CDKL5 properties; in particular, it negatively influences the catalytic activity and is required for its proper sub-nuclear localization. We propose a model in which CDKL5 phosphorylation is required for its entrance into the nucleus whereas a portion of the COOH-terminal domain is responsible for a stable residency in this cellular compartment probably through protein-protein interactions

Multiple Layers of CDK5R1 Regulation in Alzheimer’s Disease Implicate Long Non-Coding RNAs

Cyclin-dependent kinase 5 regulatory subunit 1 (CDK5R1) gene encodes for p35, the main activator of Cyclin-dependent kinase 5 (CDK5). The active p35/CDK5 complex is involved in numerous aspects of brain development and function, and its deregulation is closely associated to Alzheimer\u2019s disease (AD) onset and progression. We recently showed that miR-15/107 family can negatively regulate CDK5R1 expression modifying mRNA stability. Interestingly, miRNAs belonging to miR-15/107 family are downregulated in AD brain while CDK5R1 is upregulated. Long non-coding RNAs (lncRNAs) are emerging as master regulators of gene expression, including miRNAs, and their dysregulation has been implicated in the pathogenesis of AD. Here, we evaluated the existence of an additional layer of CDK5R1 expression regulation provided by lncRNAs. In particular, we focused on three lncRNAs potentially regulating CDK5R1 expression levels, based on existing data: NEAT1, HOTAIR, and MALAT1. We demonstrated that NEAT1 and HOTAIR negatively regulate CDK5R1 mRNA levels, while MALAT1 has a positive effect. We also showed that all three lncRNAs positively control miR-15/107 family of miRNAs. Moreover, we evaluated the expression of NEAT1, HOTAIR, and MALAT1 in AD and control brain tissues. Interestingly, NEAT1 displayed increased expression levels in temporal cortex and hippocampus of AD patients. Interestingly, we observed a strong positive correlation between CDK5R1 and NEAT1 expression levels in brain tissues, suggesting a possible neuroprotective role of NEAT1 in AD to compensate for increased CDK5R1 levels. Overall, our work provides evidence of another level of CDK5R1 expression regulation mediated by lncRNAs and points to NEAT1 as a biomarker, as well as a potential pharmacological target for AD therapy

Foot Bone in Vivo: Its Center of Mass and Centroid of Shape

This paper studies foot bone geometrical shape and its mass distribution and establishes an assessment method of bone strength. Using spiral CT scanning, with an accuracy of sub-millimeter, we analyze the data of 384 pieces of foot bones in vivo and investigate the relationship between the bone's external shape and internal structure. This analysis is explored on the bases of the bone's center of mass and its centroid of shape. We observe the phenomenon of superposition of center of mass and centroid of shape fairly precisely, indicating a possible appearance of biomechanical organism. We investigate two aspects of the geometrical shape, (i) distance between compact bone's centroid of shape and that of the bone and (ii) the mean radius of the same density bone issue relative to the bone's centroid of shape. These quantities are used to interpret the influence of different physical exercises imposed on bone strength, thereby contributing to an alternate assessment technique to bone strength.Comment: 9 pages, 4 figure