Mouse nuclear myosin I knock-out shows interchangeability and redundancy of myosin isoforms in the cell nucleus.

Nuclear myosin I (NM1) is a nuclear isoform of the well-known "cytoplasmic" Myosin 1c protein (Myo1c). Located on the 11(th) chromosome in mice, NM1 results from an alternative start of transcription of the Myo1c gene adding an extra 16 amino acids at the N-terminus. Previous studies revealed its roles in RNA Polymerase I and RNA Polymerase II transcription, chromatin remodeling, and chromosomal movements. Its nuclear localization signal is localized in the middle of the molecule and therefore directs both Myosin 1c isoforms to the nucleus. In order to trace specific functions of the NM1 isoform, we generated mice lacking the NM1 start codon without affecting the cytoplasmic Myo1c protein. Mutant mice were analyzed in a comprehensive phenotypic screen in cooperation with the German Mouse Clinic. Strikingly, no obvious phenotype related to previously described functions has been observed. However, we found minor changes in bone mineral density and the number and size of red blood cells in knock-out mice, which are most probably not related to previously described functions of NM1 in the nucleus. In Myo1c/NM1 depleted U2OS cells, the level of Pol I transcription was restored by overexpression of shRNA-resistant mouse Myo1c. Moreover, we found Myo1c interacting with Pol II. The ratio between Myo1c and NM1 proteins were similar in the nucleus and deletion of NM1 did not cause any compensatory overexpression of Myo1c protein. We observed that Myo1c can replace NM1 in its nuclear functions. Amount of both proteins is nearly equal and NM1 knock-out does not cause any compensatory overexpression of Myo1c. We therefore suggest that both isoforms can substitute each other in nuclear processes

PPAR-α and glucocorticoid receptor synergize to promote erythroid progenitor self-renewal

Many acute and chronic anaemias, including haemolysis, sepsis and genetic bone marrow failure diseases such as Diamond–Blackfan anaemia, are not treatable with erythropoietin (Epo), because the colony-forming unit erythroid progenitors (CFU-Es) that respond to Epo are either too few in number or are not sensitive enough to Epo to maintain sufficient red blood cell production. Treatment of these anaemias requires a drug that acts at an earlier stage of red cell formation and enhances the formation of Epo-sensitive CFU-E progenitors. Recently, we showed that glucocorticoids specifically stimulate self-renewal of an early erythroid progenitor, burst-forming unit erythroid (BFU-E), and increase the production of terminally differentiated erythroid cells. Here we show that activation of the peroxisome proliferator-activated receptor α (PPAR-α) by the PPAR-α agonists GW7647 and fenofibrate synergizes with the glucocorticoid receptor (GR) to promote BFU-E self-renewal. Over time these agonists greatly increase production of mature red blood cells in cultures of both mouse fetal liver BFU-Es and mobilized human adult CD34+ peripheral blood progenitors, with a new and effective culture system being used for the human cells that generates normal enucleated reticulocytes. Although Ppara−/− mice show no haematological difference from wild-type mice in both normal and phenylhydrazine (PHZ)-induced stress erythropoiesis, PPAR-α agonists facilitate recovery of wild-type but not Ppara−/− mice from PHZ-induced acute haemolytic anaemia. We also show that PPAR-α alleviates anaemia in a mouse model of chronic anaemia. Finally, both in control and corticosteroid-treated BFU-E cells, PPAR-α co-occupies many chromatin sites with GR; when activated by PPAR-α agonists, additional PPAR-α is recruited to GR-adjacent sites and presumably facilitates GR-dependent BFU-E self-renewal. Our discovery of the role of PPAR-α agonists in stimulating self-renewal of early erythroid progenitor cells suggests that the clinically tested PPAR-α agonists we used may improve the efficacy of corticosteroids in treating Epo-resistant anaemias.United States. Defense Advanced Research Projects Agency (Grant HR0011-14-2-0005)United States. Army Medical Research and Materiel Command (Grant W81WH-12-1-0449)National Heart, Lung, and Blood Institute (Grant 2 P01 HL032262-25

SBDS Expression and Localization at the Mitotic Spindle in Human Myeloid Progenitors

BACKGROUND: Shwachman-Diamond Syndrome (SDS) is a hereditary disease caused by mutations in the SBDS gene. SDS is clinically characterized by pancreatic insufficiency, skeletal abnormalities and bone marrow dysfunction. The hematologic abnormalities include neutropenia, neutrophil chemotaxis defects, and an increased risk of developing Acute Myeloid Leukemia (AML). Although several studies have suggested that SBDS as a protein plays a role in ribosome processing/maturation, its impact on human neutrophil development and function remains to be clarified. METHODOLOGY/PRINCIPAL FINDINGS: We observed that SBDS RNA and protein are expressed in the human myeloid leukemia PLB-985 cell line and in human hematopoietic progenitor cells by quantitative RT-PCR and Western blot analysis. SBDS expression is downregulated during neutrophil differentiation. Additionally, we observed that the differentiation and proliferation capacity of SDS-patient bone marrow hematopoietic progenitor cells in a liquid differentiation system was reduced as compared to control cultures. Immunofluorescence analysis showed that SBDS co-localizes with the mitotic spindle and in vitro binding studies reveal a direct interaction of SBDS with microtubules. In interphase cells a perinuclear enrichment of SBDS protein which co-localized with the microtubule organizing center (MTOC) was observed. Also, we observed that transiently expressed SDS patient-derived SBDS-K62 or SBDS-C84 mutant proteins could co-localize with the MTOC and mitotic spindle. CONCLUSIONS/SIGNIFICANCE: SBDS co-localizes with the mitotic spindle, suggesting a role for SBDS in the cell division process, which corresponds to the decreased proliferation capacity of SDS-patient bone marrow CD34(+) hematopoietic progenitor cells in our culture system and also to the neutropenia in SDS patients. A role in chromosome missegregation has not been clarified, since similar spatial and time-dependent localization is observed when patient-derived SBDS mutant proteins are studied. Thus, the increased risk of myeloid malignancy in SDS remains unexplained

Physical complaints in ageing persons with spinal muscular atrophy.

Contains fulltext : 49152.pdf (publisher's version ) (Open Access)OBJECTIVE: While life expectancy is improving for persons with spinal muscular atrophy, new physical complaints may arise. To investigate this, we studied persons with a long duration and severe course (high functional limitations) of the disease. DESIGN: Cross-sectional descriptive study. SUBJECTS/PATIENTS: Persons with spinal muscular atrophy. METHODS: Questionnaires and structured interviews on prevalence of physical complaints and their duration. Of 190 questionnaires 99 were returned; of 23 persons (with the longest disease duration and high functional limitation level) selected for structured medical interviews 9 participated. RESULTS: Patterns common within and different between the different types of spinal muscular atrophy were identified. Of the 10 most common complaints, types 1-2 had a significantly higher prevalence of kyphoscoliosis, difficulty in coughing, joint contractures and voice/speech problems, while type 3 had a significantly higher prevalence of fatigue. No statistically significant correlation was found between the appearance of physical complaints and disease duration. However, sleeping and swallowing problems were in the 5 most common complaints with the shortest mean time of appearance. The structured interview revealed hypermobility in the hand, suffusion of the eyes, and itching as new complaints with high prevalence. CONCLUSION: There are indications that the frequency of less well-known physical complaints increases with ageing

Correlation between motor performance scales, body composition, and anthropometry in patients with duchenne muscular dystrophy

The aim of this study is to investigate the relationship between body composition, anthropometry, and motor scales in patients with Duchenne muscular dystrophy (DMD). Twenty six patients with DMD were evaluated by Expanded Hammersmith Functional Motor Scale (HFMSE), gross motor function classification system (GMFCS), multifrequency bioelectrical impedance analysis, and anthropometric measurements. Seventeen healthy children served as control group. There were 26 patients with a mean age of 9.5 +/- A 4.8 years. Ages and anthropometric measurements did not differ between groups. Of the 26 patients, nine were level I, seven were level II, two were level III, seven were level IV, and one was level V, according to the GMFCS. Despite the similar percentage of total body water, extracellular water/intracellular water ratio was significantly elevated in DMD patients (p = 0.001). Increased values of fat percentage and body fat mass index (BFMI) correlated positively with elevated GMFCS levels (r = 0.785 and 0.719 respectively). Increased fat-free mass index (FFMI) correlated negatively with elevated GMFCS levels (r = -0.401). Increased fat percentage and BFMI correlated negatively with HFMSE scores (r = -0.779 and -0.698, respectively). Increased values of FFMI correlated positively with HFMSE scores. There was also a negative correlation between increased skin fold measurements from triceps and scapula and HFMSE scores (r = -0.618 and -0.683, respectively). Increased skin fold values from the same regions correlated positively with elevated GMFCS levels (r = 0.643 and 0.712, respectively). Significant body composition changes occur in patients with DMD. Anthropometric and multifrequency bioelectrical impedance analyses measurements show good correlation between motor function scales. These results may also be helpful to evaluate the effects of new treatment strategies