21 research outputs found
The Gag Cleavage Product, p12, is a Functional Constituent of the Murine Leukemia Virus Pre-Integration Complex
The p12 protein is a cleavage product of the Gag precursor of the murine leukemia virus (MLV). Specific mutations in p12 have been described that affect early stages of infection, rendering the virus replication-defective. Such mutants showed normal generation of genomic DNA but no formation of circular forms, which are markers of nuclear entry by the viral DNA. This suggested that p12 may function in early stages of infection but the precise mechanism of p12 action is not known. To address the function and follow the intracellular localization of the wt p12 protein, we generated tagged p12 proteins in the context of a replication-competent virus, which allowed for the detection of p12 at early stages of infection by immunofluorescence. p12 was found to be distributed to discrete puncta, indicative of macromolecular complexes. These complexes were localized to the cytoplasm early after infection, and thereafter accumulated adjacent to mitotic chromosomes. This chromosomal accumulation was impaired for p12 proteins with a mutation that rendered the virus integration-defective. Immunofluorescence demonstrated that intracellular p12 complexes co-localized with capsid, a known constituent of the MLV pre-integration complex (PIC), and immunofluorescence combined with fluorescent in situ hybridization (FISH) revealed co-localization of the p12 proteins with the incoming reverse transcribed viral DNA. Interactions of p12 with the capsid and with the viral DNA were also demonstrated by co-immunoprecipitation. These results imply that p12 proteins are components of the MLV PIC. Furthermore, a large excess of wt PICs did not rescue the defect in integration of PICs derived from mutant p12 particles, demonstrating that p12 exerts its function as part of this complex. Altogether, these results imply that p12 proteins are constituent of the MLV PIC and function in directing the PIC from the cytoplasm towards integration
HIV infection of non-dividing cells: a divisive problem
Understanding how lentiviruses can infect terminally differentiated, non-dividing cells has proven a very complex and controversial problem. It is, however, a problem worth investigating, for it is central to HIV-1 transmission and AIDS pathogenesis. Here I shall attempt to summarise what is our current understanding for HIV-1 infection of non-dividing cells. In some cases I shall also attempt to make sense of controversies in the field and advance one or two modest proposals
Sucrose, But Not Glucose, Blocks IL1-β-Induced Inflammatory Response in Human Chondrocytes by Inducing Autophagy via AKT/mTOR Pathway
Human Immunodeficiency Virus Type 1 Central DNA Flap: Dynamic Terminal Product of Plus-Strand Displacement DNA Synthesis Catalyzed by Reverse Transcriptase Assisted by Nucleocapsid Protein
To terminate the reverse transcription of the human immunodeficiency virus type 1 (HIV-1) genome, a final step occurs within the center of the proviral DNA generating a 99-nucleotide DNA flap (6). This step, catalyzed by reverse transcriptase (RT), is defined as a discrete strand displacement (SD) synthesis between the first nucleotide after the central priming (cPPT) site and the final position of the central termination sequence (CTS) site. Using recombinant HIV-1 RT and a circular single-stranded DNA template harboring the cPPT-CTS sequence, we have developed an SD synthesis-directed in vitro termination assay. Elongation, strand displacement, and complete central flap behavior were analyzed using electrophoresis and electron microscopy approaches. Optimal conditions to obtain complete central flap, which ended at the CTS site, have been defined in using nucleocapsid protein (NCp), the main accessory protein of the reverse transcription complex. A full-length HIV-1 central DNA flap was then carried out in vitro. Its synthesis appears faster in the presence of the HIV-1 NCp or the T4-encoded SSB protein (gp32). Finally, a high frequency of strand transfer was shown during the SD synthesis along the cPPT-CTS site with RT alone. This reveals a local and efficient 3′-5′ branch migration which emphasizes some important structural fluctuations within the flap. These fluctuations may be stabilized by the NCp chaperone activity. The biological implications of the RT-directed NCp-assisted flap synthesis are discussed within the context of reverse transcription complexes, assembly of the preintegration complexes, and nuclear import of the HIV-1 proviral DNA to the nucleus toward their chromatin targets
Growth Charts for Individuals with Mucopolysaccharidosis VI (Maroteaux–Lamy Syndrome)
Background: The skeletal phenotype of mucopolysaccharidosis VI (MPS VI) is characterized by short stature and growth failure. Objective: The purpose of this study was to construct reference growth curves for MPS VI patients with rapidly and slowly progressive disease. Methods: We pooled cross-sectional and longitudinal height for age data from galsulfase (Naglazyme(®), BioMarin Pharmaceutical Inc.), treatment naïve patients (n = 269) who participated in various MPS VI studies, including galsulfase clinical trials and their extension programs, the MPS VI clinical surveillance program (CSP), and the MPS VI survey and resurvey studies, to construct growth charts for the MPS VI population. There were 229 patients included in this study, of which data from 207 patients ≤25 years of age with 513 height measurements were used for constructing reference growth curves. Results: Height for age growth curves for the 5th, 10th, 25th, 50th, 75th, 90th, and 95th percentiles were constructed for patients with rapidly and slowly progressing disease defined by the pre-enzyme replacement therapy (ERT) uGAG levels of > or ≤200 μg/mg creatinine. The mean (SD) pre-ERT uGAG levels were 481.0 (218.6) and 97.8 (56.3) μg/mg creatinine for the patients ≤25 years of age with rapidly (n = 131) and slowly (n = 76) progressing MPS VI disease, respectively. The median growth curves for patients with ≤ and >200 μg/mg creatinine were above and below the median (50th percentile) growth curve for the entire MPS VI population. Conclusion: MPS VI growth charts have been developed to assist in the clinical management of MPS VI patients. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this chapter (doi:10.1007/8904_2014_333) contains supplementary material, which is available to authorized users
Misfolding of vasopressin receptors: biased agonist pharmacochaperones as potential therapeutics
International audienceBiased agonists and pharmacological chaperones have demonstrated their potential to harness G protein-coupled receptor signaling and trafficking, and have collectively opened new possibilities in G protein-coupled receptor drug discovery. Combining pharmacological chaperoning and biased agonism properties into a unique given molecule would be of high therapeutic interest in many human diseases resulting from G protein-coupled receptor mutation and misfolding. This strategy perfectly fits to congenital Nephrogenic Diabetes Insipidus which is a typical conformational disease. In most of the cases, it is associated to inactivating mutations of the renal arginine-vasopressin V2 receptor leading to misfolding and intracellular retention of the receptor, causing the inability of patients to concentrate their urine in response to the antidiuretic hormone. Cell-permeable pharmacological chaperones have been successfully challenged to restore plasma membrane localization of the receptor mutants and to rescue their function. Interestingly, different classes of pharmacological chaperones of the V2 receptor have proven their usefulness and efficacy, such as antagonists, agonists as well as biased agonists. These compounds, particularly small-molecule biased agonists which elicit the V2-induced Gs protein-dependent signaling pathway, but not V2-related arrestin-dependent cell responses, represent a potential therapeutic treatment of this X-linked genetic pathology
Management of Pediatric Acute Hematogenous Osteomyelitis, Part II: A Focus on Methicillin-Resistant Staphylococcus aureus
The PH domain proteins IPIP27A and B link OCRL1 to receptor recycling in the endocytic pathway
We identify two new binding partners for the OCRL1 protein that is mutated in Lowe syndrome and type 2 Dent disease, which we call IPIP27A and B. The IPIPs are required for receptor recycling in the endocytic pathway, suggesting that defects in this process lead to the aforementioned disorders