Limb-Girdle Muscular Dystrophy Type 2C: Case Report

Limb-Girdle muscular dystrophy (LGMD) is a group of inherited disorders that lead to muscle weakness and skeletal muscle wasting involving the muscles around the hips and shoulders. This can cause a gait disturbance, difficulty running or even a complete loss of the ability to walk. The appearance of the disorder, the course and the muscles affected are variable between the different subtypes of the disease. Typically, patients with type 2C Limb-Girdle dystrophy (LGMD2C) start having symptoms early in infancy and lose their ability to walk around the age of 12. Others have less symptomatology and have late expression in adulthood. This disorder can affect the heart muscle in some patients. They can also have osteoarticular and vertebral deformations. The prognosis depends on the muscles often affected by respiratory failure.LGMD2C is caused by a pathogenic mutation in the SGCG gene. We report the case of a 13-year-old child, with a notion of femoral fracture at the age of 5 years and first degree consanguinity in the parents, no similar case in the family, and who presents since 3 years difficulty walking Clinical examination: walking, positive sign of Gowers, scapula alta, enlarged calves. On the biological level: CPK 6380, LDH 482, PAL 219, ASAT 68, ALAT 103. The electromyogram shows a slowing of the motor conduction speed of the external popliteal sciatic nerve with a slight loss of amplitude. Muscle biopsy objective a discreet dystrophic formula with a total absence of expression of gamma sarcoglycans and proteins of the muscle membrane. Gamma glycanopathy is genetically confirmed by the mutation of the δ-SG gene. A cardiac ultrasound was without abnormality.

Identification and Characterization of the Nucleolar Localization Signal of Autographa californica Multiple Nucleopolyhedrovirus LEF5

ABSTRACT Autographa californica multiple nucleopolyhedrovirus (AcMNPV) late expression factor 5 (LEF5) is highly conserved in all sequenced baculovirus genomes and plays an important role in production of infectious viral progeny. In this study, nucleolar localization of AcMNPV LEF5 was characterized. Through transcriptome analysis, we identified two putative nucleolar proteins, Spodoptera frugiperda nucleostemin (SfNS) and fibrillarin (SfFBL), from Sf9 cells. Immunofluorescence analysis demonstrated that SfNS and SfFBL were localized to the nucleolus. AcMNPV infection resulted in reorganization of the nucleoli of infected cells. Colocalization of LEF5 and SfNS showed that AcMNPV LEF5 was localized to the nucleolus in Sf9 cells. Bioinformatic analysis revealed that basic amino acids of LEF5 are enriched at residues 184 to 213 and may contain a nucleolar localization signal (NoLS). Green fluorescent protein (GFP) fused to NoLS of AcMNPV LEF5 localized to the nucleoli of transfected cells. Multiple-point mutation analysis demonstrated that amino acid residues 197 to 204 are important for nucleolar localization of LEF5. To identify whether the NoLS in AcMNPV LEF5 is important for production of viral progeny, a lef5-null AcMNPV bacmid was constructed; several NoLS-mutated LEF5 proteins were reinserted into the lef5-null AcMNPV bacmid with a GFP reporter. The constructs containing point mutations at residues 185 to 189 or 197 to 204 in AcMNPV LEF5 resulted in reduction in production of infectious viral progeny and occlusion body yield in bacmid-transfected cells. Together, these data suggested that AcMNPV LEF5 contains an NoLS, which is important for nucleolar localization of LEF5, progeny production, and occlusion body production. IMPORTANCE Many viruses, including human and plant viruses, target nucleolar functions as part of their infection strategy. However, nucleolar localization for baculovirus proteins has not yet been characterized. In this study, two nucleolar proteins, SfNS and SfFBL, were identified in Sf9 cells. Our results showed that Autographa californica multiple nucleopolyhedrovirus (AcMNPV) infection resulted in redistribution of the nucleoli of infected cells. We demonstrated that AcMNPV late expression factor 5 (LEF5) could localize to the nucleolus and contains a nucleolar localization signal (NoLS), which is important for nucleolar localization of AcMNPV LEF5 and for production of viral progeny and yield of occlusion bodies.

STATs in Lung Development: Distinct Early and Late Expression, Growth Modulation and Signaling Dysregulation in Congenital Diaphragmatic Hernia

Background: Congenital diaphragmatic hernia (CDH) is a life-threatening developmental anomaly, intrinsically combining severe pulmonary hypoplasia and hypertension. During development, signal transducers and activators of transcription (STAT) are utilized to elicit cell growth, differentiation, and survival. Methods: We used the nitrofen-induced CDH rat model. At selected gestational time points, lungs were divided into two experimental groups, i.e., control or CDH. We performed immunohistochemistry and western blotting analysis to investigate the developmental expression profile of the complete family of STATs (STAT1-6), plus specific STATs activation (p-STAT3, p-STAT6) and regulation by SOCS (SOCS3) in normal lungs against those of diseased lungs. The normal fetal lung explants were treated with piceatannol (STAT3 inhibitor) in vitro followed by morphometrical analysis. Results: Molecular profiling of STATs during the lung development revealed distinct early and late expression signatures. Experimental CDH altered the STATs expression, activation, and regulation in the fetal lungs. In particular, STAT3 and STAT6 were persistently over-expressed and early over-activated. Piceatannol treatment dose-dependently stimulated the fetal lung growth. Conclusion: These findings suggest that STATs play an important role during normal fetal lung development and CDH pathogenesis. Moreover, functionally targeting STAT signaling modulates fetal lung growth, which highlights that STAT3 and STAT6 signaling might be promising therapeutic targets in reducing or preventing pulmonary hypoplasia in CDH.

Childhood tolerance of severe influenza: a mortality analysis in mice

During the 1918 influenza pandemic, children experienced substantially lower mortality than adults, a striking but unexplained finding. Whether this was due to enhanced resistance (reduced virus load) or better tolerance (reduced impact of infection) has not been defined. We found that prepubertal mice infected with H1N1 influenza virus also showed greater survival than infected pubertal mice, despite similar virus loads. Transcriptome profiling of infected lungs identified estrogen as a regulator of susceptibility in both sexes and also linked better survival to late expression of IL-1β. Blocking puberty with gonadectomy or a gonadotropin-releasing hormone antagonist improved survival. Estrogen or testosterone (which can be converted to estrogen) restored susceptibility of gonadectomized pubertal mice to influenza mortality, but dihydrotestosterone (which cannot be converted to estrogen) did not. Estrogen receptor blockade with fulvestrant in both male and female pubertal mice resulted in improved survival, even when given 3 days after infection. Moreover, late, but not early, IL-1β neutralization after infection was also protective. These findings indicate that pubertal increases in estrogen in both sexes are associated with increased mortality during influenza. This helps explain the reduced mortality of children seen with influenza in 1918 and might also be relevant to childhood tolerance to many other infectious diseases.

Importance of codon usage for the temporal regulation of viral gene expression

The glycoproteins of herpesviruses and of HIV/SIV are made late in the replication cycle and are derived from transcripts that use an unusual codon usage that is quite different from that of the host cell. Here we show that the actions of natural transinducers from these two different families of persistent viruses (Rev of SIV and ORF57 of the rhesus monkey rhadinovirus) are dependent on the nature of the skewed codon usage. In fact, the transinducibility of expression of these glycoproteins by Rev and by ORF57 can be flipped simply by changing the nature of the codon usage. Even expression of a luciferase reporter could be made Rev dependent or ORF57 dependent by distinctive changes to its codon usage. Our findings point to a new general principle in which different families of persisting viruses use a poor codon usage that is skewed in a distinctive way to temporally regulate late expression of structural gene products.

Characterization of protein–protein interaction domains within the baculovirus Autographa californica multiple nucleopolyhedrovirus late expression factor LEF-3

Autographa californica nucleopolyhedrovirus late expression factor 3 (LEF-3) is required for late viral gene expression probably through its numerous functions related to DNA replication, including nuclear localization of the virus helicase P143 and binding to ssDNA. LEF-3 appears to interact with itself as a homo-oligomer, although the details of this oligomeric structure are not yet known. To examine LEF-3–LEF-3 interactions, a bimolecular fluorescent protein complementation assay was used. Pairs of recombinant plasmids expressing full-length LEF-3 fused to one of two complementary fragments (V1 or V2) of a variant of yellow fluorescent protein named ‘Venus’ were constructed. Plasmids expressing fusions with complementary fragments of Venus were co-transfected into Sf21 cells and analysed by fluorescence microscopy. Co-transfected plasmids expressing full-length V1–LEF-3 and V2–LEF-3 showed positive fluorescence, confirming the formation of homo-oligomers. A series of truncated V1/V2–LEF-3 fusions was constructed and used to investigate interactions with one another as well as with full-length LEF-3.