Protein multi-scale organization through graph partitioning and robustness analysis: Application to the myosin-myosin light chain interaction

Despite the recognized importance of the multi-scale spatio-temporal organization of proteins, most computational tools can only access a limited spectrum of time and spatial scales, thereby ignoring the effects on protein behavior of the intricate coupling between the different scales. Starting from a physico-chemical atomistic network of interactions that encodes the structure of the protein, we introduce a methodology based on multi-scale graph partitioning that can uncover partitions and levels of organization of proteins that span the whole range of scales, revealing biological features occurring at different levels of organization and tracking their effect across scales. Additionally, we introduce a measure of robustness to quantify the relevance of the partitions through the generation of biochemically-motivated surrogate random graph models. We apply the method to four distinct conformations of myosin tail interacting protein, a protein from the molecular motor of the malaria parasite, and study properties that have been experimentally addressed such as the closing mechanism, the presence of conserved clusters, and the identification through computational mutational analysis of key residues for binding.Comment: 13 pages, 7 Postscript figure

Reproducibility of Ablated Volume Measurement Is Higher with Contrast-Enhanced Ultrasound than with B-Mode Ultrasound after Benign Thyroid Nodule Radiofrequency Ablation—A Preliminary Study

The reproducibility of contrast-enhanced ultrasound (CEUS) and standard B-mode ultrasound in the assessment of radiofrequency-ablated volume of benign thyroid nodules was compared. A preliminary study was conducted on consecutive patients who underwent radiofrequency ablation (RFA) of benign thyroid nodules between 2014 and 2016, with available CEUS and B-mode post-ablation checks. CEUS and B-mode images were retrospectively evaluated by two radiologists to assess inter- and intra-observer agreement in the assessment of ablated volume (Bland-Altman test). For CEUS, the mean inter-observer difference (95% limits of agreement) was 0.219 mL (-0.372-0.809 mL); for B-mode, the mean difference was 0.880 mL (-1.655-3.414 mL). Reproducibility was significantly higher for CEUS (85%) than for B-mode (27%). Mean intra-observer differences (95% limits of agreement) were 0.013 mL (0.803-4.097 mL) for Reader 1 and 0.031 mL (0.763-3.931 mL) for Reader 2 using CEUS, while they were 0.567 mL (-2.180-4.317 mL, Reader 1) and 0.759 mL (-2.584-4.290 mL, Reader 2) for B-mode. Intra-observer reproducibility was significantly higher for CEUS (96% and 95%, for the two readers) than for B-mode (21% and 23%). In conclusion, CEUS had higher reproducibility and inter- and intra-observer agreement compared to conventional B-mode in the assessment of radiofrequency-ablated volume of benign thyroid nodules

The ocular albinism type 1 protein, an intracellular G protein-coupled receptor, regulates melanosome transport in pigment cells

The protein product of the ocular albinism type 1 gene, named OA1, is a pigment cell-specific G protein-coupled receptor exclusively localized to intracellular organelles, namely lysosomes and melanosomes. Loss of OA1 function leads to the formation of macromelanosomes, suggesting that this receptor is implicated in organelle biogenesis, however the mechanism involved in the pathogenesis of the disease remains obscure. We report here the identification of an unexpected abnormality in melanosome distribution both in retinal pigment epithelium (RPE) and skin melanocytes of Oa1-knock-out (KO) mice, consisting in a displacement of the organelles from the central cytoplasm towards the cell periphery. Despite their depletion from the microtubule (MT)-enriched perinuclear region, Oa1-KO melanosomes were able to aggregate at the centrosome upon disruption of the actin cytoskeleton or expression of a dominant-negative construct of myosin Va. Consistently, quantification of organelle transport in living cells revealed that Oa1-KO melanosomes displayed a severe reduction in MT-based motility; however, this defect was rescued to normal following inhibition of actin-dependent capture at the cell periphery. Together, these data point to a defective regulation of organelle transport in the absence of OA1 and imply that the cytoskeleton might represent a downstream effector of this receptor. Furthermore, our results enlighten a novel function for OA1 in pigment cells and suggest that ocular albinism type 1 might result from a different pathogenetic mechanism than previously thought, based on an organelle-autonomous signalling pathway implicated in the regulation of both membrane traffic and transport

Eight previously unidentified mutations found in the OA1 ocular albinism gene

BACKGROUND: Ocular albinism type 1 (OA1) is an X-linked ocular disorder characterized by a severe reduction in visual acuity, nystagmus, hypopigmentation of the retinal pigmented epithelium, foveal hypoplasia, macromelanosomes in pigmented skin and eye cells, and misrouting of the optical tracts. This disease is primarily caused by mutations in the OA1 gene. METHODS: The ophthalmologic phenotype of the patients and their family members was characterized. We screened for mutations in the OA1 gene by direct sequencing of the nine PCR-amplified exons, and for genomic deletions by PCR-amplification of large DNA fragments. RESULTS: We sequenced the nine exons of the OA1 gene in 72 individuals and found ten different mutations in seven unrelated families and three sporadic cases. The ten mutations include an amino acid substitution and a premature stop codon previously reported by our team, and eight previously unidentified mutations: three amino acid substitutions, a duplication, a deletion, an insertion and two splice-site mutations. The use of a novel Taq polymerase enabled us to amplify large genomic fragments covering the OA1 gene. and to detect very likely six distinct large deletions. Furthermore, we were able to confirm that there was no deletion in twenty one patients where no mutation had been found. CONCLUSION: The identified mutations affect highly conserved amino acids, cause frameshifts or alternative splicing, thus affecting folding of the OA1 G protein coupled receptor, interactions of OA1 with its G protein and/or binding with its ligand

Developmental regulation of MURF E3 ubiquitin ligases in skeletal muscle

The striated muscle-specific tripartite motif (TRIM) proteins TRIM63/MURF1, TRIM55/MURF2 and TRIM54/MURF3 can function as E3 ubiquitin ligases in ubiquitin-mediated muscle protein turnover. Despite the well-characterised role of MURF1 in skeletal muscle atrophy, the dynamics of MURF isogene expression in the development and early postnatal adaptation of skeletal muscle is unknown. Here, we show that MURF2 is the isogene most highly expressed in embryonic skeletal muscle at E15.5, with the 50 kDa A isoform predominantly expressed. MURF1 and MURF3 are upregulated only postnatally. Knockdown of MURF2 p50A by isoform-specific siRNA results in delayed myogenic differentiation and myotube formation in vitro, with perturbation of the stable, glutamylated microtubule population. This underscores that MURF2 plays an important role in the earliest stages of skeletal muscle differentiation and myofibrillogenesis. During further development, there is a shift towards the 60 kDa A isoform, which dominates postnatally. Analysis of the fibre-type expression shows that MURF2 A isoforms are predominantly slow-fibre associated, whilst MURF1 is largely excluded from these fibres, and MURF3 is ubiquitously distributed in both type I and II fibres

Developing cardiac and skeletal muscle share fast-skeletal myosin heavy chain and cardiac troponin-I expression

Skeletal muscle derived stem cells (MDSCs) transplanted into injured myocardium can differentiate into fast skeletal muscle specific myosin heavy chain (sk-fMHC) and cardiac specific troponin-I (cTn-I) positive cells sustaining recipient myocardial function. We have recently found that MDSCs differentiate into a cardiomyocyte phenotype within a three-dimensional gel bioreactor. It is generally accepted that terminally differentiated myocardium or skeletal muscle only express cTn-I or sk-fMHC, respectively. Studies have shown the presence of non-cardiac muscle proteins in the developing myocardium or cardiac proteins in pathological skeletal muscle. In the current study, we tested the hypothesis that normal developing myocardium and skeletal muscle transiently share both sk-fMHC and cTn-I proteins. Immunohistochemistry, western blot, and RT-PCR analyses were carried out in embryonic day 13 (ED13) and 20 (ED20), neonatal day 0 (ND0) and 4 (ND4), postnatal day 10 (PND10), and 8 week-old adult female Lewis rat ventricular myocardium and gastrocnemius muscle. Confocal laser microscopy revealed that sk-fMHC was expressed as a typical striated muscle pattern within ED13 ventricular myocardium, and the striated sk-fMHC expression was lost by ND4 and became negative in adult myocardium. cTn-I was not expressed as a typical striated muscle pattern throughout the myocardium until PND10. Western blot and RT-PCR analyses revealed that gene and protein expression patterns of cardiac and skeletal muscle transcription factors and sk-fMHC within ventricular myocardium and skeletal muscle were similar at ED20, and the expression patterns became cardiac or skeletal muscle specific during postnatal development. These findings provide new insight into cardiac muscle development and highlight previously unknown common developmental features of cardiac and skeletal muscle. © 2012 Clause et al

Prenatal muscle development in a mouse model for the secondary dystroglycanopathies

The defective glycosylation of α-dystroglycan is associated with a group of muscular dystrophies that are collectively referred to as the secondary dystroglycanopathies. Mutations in the gene encoding fukutin-related protein (FKRP) are one of the most common causes of secondary dystroglycanopathy in the UK and are associated with a wide spectrum of disease. Whilst central nervous system involvement has a prenatal onset, no studies have addressed prenatal muscle development in any of the mouse models for this group of diseases. In view of the pivotal role of α-dystroglycan in early basement membrane formation, we sought to determine if the muscle formation was altered in a mouse model of FKRP-related dystrophy

Expression of the myosin heavy chain IIB gene in porcine skeletal muscle: the role of the CArG-box promoter response element

Due to its similarity to humans, the pig is increasingly being considered as a good animal model for studying a range of human diseases. Despite their physiological similarities, differential expression of the myosin heavy chain (MyHC) IIB gene (MYH4) exists in the skeletal muscles of these species, which is associated with a different muscle phenotype. The expression of different MyHC isoforms is a critical determinant of the contractile and metabolic characteristics of the muscle fibre. We aimed to elucidate whether a genomic mechanism was responsible for the drastically different expression of MYH4 between pigs and humans, thus improving our understanding of the pig as a model for human skeletal muscle research. We utilized approximately 1 kb of the MYH4 promoter from a domestic pig and a human (which do and do not express MYH4, respectively) to elucidate the role of the promoter sequence in regulating the high expression of MYH4 in porcine skeletal muscle. We identified a 3 bp genomic difference within the proximal CArG and Ebox region of the MYH4 promoter of pigs and humans that dictates the differential activity of these promoters during myogenesis. Subtle species-specific genomic differences within the CArG-box region caused differential protein-DNA interactions at this site and is likely accountable for the differential MYH4 promoter activity between pigs and humans. We propose that the genomic differences identified herein explain the differential activity of the MYH4 promoter of pigs and humans, which may contribute to the differential expression patterns displayed in these otherwise physiologically similar mammals. Further, we report that both the pig and human MYH4 promoters can be induced by MyoD over- expression, but the capacity to activate the MYH4 promoter is largely influenced by the 3 bp difference located within the CArG-box region of the proximal MYH4 promoter

The Northern Cross Fast Radio Burst project - II. Monitoring of repeating FRB 20180916B, 20181030A, 20200120E and 20201124A

In this work we report the results of a nineteen-month Fast Radio Burst observational campaign carried out with the North-South arm of the Medicina Northern Cross radio telescope at 408~MHz in which we monitored four repeating sources: FRB20180916B, FRB20181030A, FRB20200120E and FRB20201124A. We present the current state of the instrument and the detection and characterisation of three bursts from FRB20180916B. Given our observing time, our detections are consistent with the event number we expect from the known burst rate ($2.7 \pm 1.9$ above our 10$\sigma$, 38~Jy~ms detection threshold) in the 5.2 day active window of the source, further confirming the source periodicity. We detect no bursts from the other sources. We turn this result into a 95\% confidence level lower limit on the slope of the differential fluence distribution $\alpha$ to be $\alpha > 2.1$ and $\alpha > 2.2$ for FRB20181030A and FRB20200120E respectively. Given the known rate for FRB20201124A, we expect $1.0 \pm 1.1$ bursts from our campaign, consistent with our non-detection.Comment: MNRAS Accepted, 10 pages, 6 figure

Breast imaging and cancer diagnosis during the COVID-19 pandemic: recommendations from the Italian College of Breast Radiologists by SIRM

The Italian College of Breast Radiologists by the Italian Society of Medical Radiology (SIRM) provides recommendations for breast care provision and procedural prioritization during COVID-19 pandemic, being aware that medical decisions must be currently taken balancing patient\u2019s individual and community safety: (1) patients having a scheduled or to-be-scheduled appointment for in-depth diagnostic breast imaging or needle biopsy should confirm the appointment or obtain a new one; (2) patients who have suspicious symptoms of breast cancer (in particular: new onset palpable nodule; skin or nipple retraction; orange peel skin; unilateral secretion from the nipple) should request non-deferrable tests at radiology services; (3) asymptomatic women performing annual mammographic follow-up after breast cancer treatment should preferably schedule the appointment within 1\ua0year and 3\ua0months from the previous check, compatibly with the local organizational conditions; (4) asymptomatic women who have not responded to the invitation for screening mammography after the onset of the pandemic or have been informed of the suspension of the screening activity should schedule the check preferably within 3\ua0months from the date of the not performed check, compatibly with local organizational conditions. The Italian College of Breast Radiologists by SIRM recommends precautions to protect both patients and healthcare workers (radiologists, radiographers, nurses, and reception staff) from infection or disease spread on the occasion of breast imaging procedures, particularly mammography, breast ultrasound, breast magnetic resonance imaging, and breast intervention procedures