Structural basis for Mis18 complex assembly and its implications for centromere maintenance.

The centromere, defined by the enrichment of CENP-A (a Histone H3 variant) containing nucleosomes, is a specialised chromosomal locus that acts as a microtubule attachment site. To preserve centromere identity, CENP-A levels must be maintained through active CENP-A loading during the cell cycle. A central player mediating this process is the Mis18 complex (Mis18α, Mis18ß and Mis18BP1), which recruits the CENP-A-specific chaperone HJURP to centromeres for CENP-A deposition. Here, using a multi-pronged approach, we characterise the structure of the Mis18 complex and show that multiple hetero- and homo-oligomeric interfaces facilitate the hetero-octameric Mis18 complex assembly composed of 4 Mis18α, 2 Mis18ß and 2 Mis18BP1. Evaluation of structure-guided/separation-of-function mutants reveals structural determinants essential for cell cycle controlled Mis18 complex assembly and centromere maintenance. Our results provide new mechanistic insights on centromere maintenance, highlighting that while Mis18α can associate with centromeres and deposit CENP-A independently of Mis18ß, the latter is indispensable for the optimal level of CENP-A loading required for preserving the centromere identity.

Study for the validation of the FeetMe® integrated sensor insole system compared to GAITRite® system to assess gait characteristics in patients with multiple sclerosis.

OBJECTIVE: To determine the concordance and statistical precision in gait velocity in people with multiple sclerosis (pwMS), measured with FeetMe® (insoles with pressure and motion sensors) compared with GAITRite® (classic reference system of gait analysis) in the timed 25-Feet Walk test (T25WT). METHODS: This observational, cross-sectional, prospective, single center study was conducted between September-2018 and April-2019 in pwMS aged 18-55 years, with Expanded Disability Status Scale (EDSS) 0-6.5 and relapse free ≥30 days at baseline. Primary endpoint was gait velocity. Secondary endpoints were ambulation time, cadence, and stride length assessment, while the correlation between gait variables and the clinical parameters of MS subjects was assessed as an exploratory endpoint. RESULTS: A total of 207 MS subjects were enrolled, of whom, 205 were considered in primary analysis. Most subjects were women (66.8%) and had relapsing-remitting MS (RRMS) (82.9%), with overall mean (standard deviation [SD]) age of 41.5 (8.0) year and EDSS 3.1 (2.0). There was a statistically significant (p<0.0001) and strong agreement (intra-class correlation coefficient (ICC) >0.830) in gait velocity, ambulation time and cadence assessment between FeetMe® and GAITRite®. CONCLUSIONS: Agreement between devices was strong (ICC≥0.800). FeetMe® is the first validated wearable medical device that allows gait monitoring in MS subjects, being potentially able to assess disease activity, progression, and treatment response.

Interrogation of the contribution of (endo)lysin domains to tune their bacteriolytic efficiency provides a novel clue to design superior antibacterials.

Bacteriophage-derived endolysins and bacterial autolysins (hereinafter lysins) represent a completely new class of efficient antibacterials. They prevent the development of bacterial resistance and help protect commensal microbiota, producing cell wall lysis. Here we have investigated whether the acquisition of enzymatic active domains (EADs) and cell wall binding domains (CWBDs) of balancing efficiencies could be a way of tuning natural lysin activity. The concept was applied to produce a chimeric lysin of superior antibacterial capacity using the endolysin Skl and the major pneumococcal autolysin LytA. Combination of the Skl EAD and the cell wall choline-binding domain (CBD) of LytA in the chimera QSLA increased the bacterial killing by 2 logs or more compared to parental enzymes at an equal concentration and extended the substrate range to resistant and emergent pneumococci and other pathogens of the mitis group. Contrarily, QLAS, containing LytA EAD and Skl CBD, was inactive against all tested strains, although domain structures were preserved and hydrolysis of purified cell walls maintained in both chimeras. As a whole, our study provides a novel clue to design superior lysins to fight multidrug-resistant pathogens based on domain selection, and a powerful in-vivo active lysin (QSLA) with promising therapeutic perspectives.

Structural and Functional Insights Into Skl and Pal Endolysins, Two Cysteine-Amidases With Anti-pneumococcal Activity. Dithiothreitol (DTT) Effect on Lytic Activity.

We have structurally and functionally characterized Skl and Pal endolysins, the latter being the first endolysin shown to kill effectively Streptococcus pneumoniae, a leading cause of deathly diseases. We have proved that Skl and Pal are cysteine-amidases whose catalytic domains, from CHAP and Amidase_5 families, respectively, share an α3ß6-fold with papain-like topology. Catalytic triads are identified (for the first time in Amidase_5 family), and residues relevant for substrate binding and catalysis inferred from in silico models, including a calcium-binding site accounting for Skl dependence on this cation for activity. Both endolysins contain a choline-binding domain (CBD) with a ß-solenoid fold (homology modeled) and six conserved choline-binding loci whose saturation induced dimerization. Remarkably, Pal and Skl dimers display a common overall architecture, preserved in choline-bound dimers of pneumococcal lysins with other catalytic domains and bond specificities, as disclosed using small angle X-ray scattering (SAXS). Additionally, Skl is proved to be an efficient anti-pneumococcal agent that kills multi-resistant strains and clinical emergent-serotype isolates. Interestingly, Skl and Pal time-courses of pneumococcal lysis were sigmoidal, which might denote a limited access of both endolysins to target bonds at first stages of lysis. Furthermore, their DTT-mediated activation, of relevance for other cysteine-peptidases, cannot be solely ascribed to reversal of catalytic-cysteine oxidation.

Meiotic chromosome synapsis depends on multivalent SYCE1-SIX6OS1 interactions that are disrupted in cases of human infertility.

Meiotic reductional division depends on the synaptonemal complex (SC), a supramolecular protein assembly that mediates homologous chromosomes synapsis and promotes crossover formation. The mammalian SC has eight structural components, including SYCE1, the only central element protein with known causative mutations in human infertility. We combine mouse genetics, cellular, and biochemical studies to reveal that SYCE1 undergoes multivalent interactions with SC component SIX6OS1. The N terminus of SIX6OS1 binds and disrupts SYCE1's core dimeric structure to form a 1:1 complex, while their downstream sequences provide a distinct second interface. These interfaces are separately disrupted by SYCE1 mutations associated with nonobstructive azoospermia and premature ovarian failure (POF), respectively. Mice harboring SYCE1's POF mutation and a targeted deletion within SIX6OS1's N terminus are infertile with failure of chromosome synapsis. We conclude that both SYCE1-SIX6OS1 binding interfaces are essential for SC assembly, thus explaining how SYCE1's reported clinical mutations give rise to human infertility.

Pancreatoblastoma associated with incomplete Beckwith-Wiedemann syndrome: case report and review of the literature.

A case of pancreatoblastoma (PB) in a 2 month-old male infant with incomplete Beckwith-Wiedemann syndrome is presented. Clinical examination disclosed left hemihypertrophy, macroglossia, bilateral exophthalmos, and enlargement of the left testis. Imaging with ultrasound and computed tomography scan showed a well-defined, heterogeneous, and grossly cystic mass arising from the head of the pancreas. Serum alpha-fetoprotein (AFP) level was elevated. The tumor was completely resected, and the histological analysis showed PB. The patient's recovery was uneventful, and AFP returned to normal values after surgery. The child has been disease-free for 5 years, and his serum AFP remained within normal values. Six other examples of this association, PB, and Beckwith-Wiedemann syndrome are recorded in the literature. The risk of developing tumor in this syndrome (complete and incomplete form) increases when hemihypertrophy is present, and the need for routine screening examination is warranted. Beckwith-Wiedemann syndrome was suggested to be a favorable biological marker for survival in children who have intraabdominal tumors.

[Repercussion on respiratory and hemodynamic parameters with a closed system of aspiration of secretion]. / Repercusión sobre parámetros respiratorios y hemodinámicos con un sistema cerrado de aspiración de secreciones.

Aspiration of bronchial secretions is a usual technique that may have an affect on hemodynamic and respiratory parameters of the patient. Our objects has been to assess if there are changes in these parameters based on two different aspiration systems: closed (CS) or open (OS) and to also compare the times used in the process. A clinical trial was performed using the crossing over method in which aspirations were performed to the same patient with the two systems. The onset system was randomized and, after a wash-out period of 3 hours, an alternative system was established. We recorded ventilatory, gasometric (baseline and at five minutes of finishing the technique) and hemodynamic (baseline, during the procedure and at five minutes) variables. The time used in each procedure was recorded. The aspiration was always performed with preoxygenation at 100% during one minute. A total of 26 patients subjected to mechanical ventilation in the assisted/controlled way entered the study and 52 aspirations were studied. We analyzed the data with the Student's t test for paired samples and ANOVA. There were no differences in the comparisons between the different determinations for the hemodynamic and gasometric variables. In the ventilatory ones, we only found a significant increase in the respiratory frequency posterior to the OS in regards to the baseline of the same system (p = 0.016). The time used in the technique was greater for the OS (p < 0.001). It can be concluded from the results that: 1. The aspiration technique does not produce clinically important alterations in the parameters studied. 2. There are no differences between the two aspiration systems. 3. The technique with CS is faster.

Acute torsion of the spleen: diagnosis and management.

We present a 2-year-old girl with a 24-hour history of abdominal pain, fever, and vomiting. The diagnosis of acute splenic torsion was made by means of color and power Doppler ultrasound. Management of this rare surgical emergency is discussed.