DNA recognition properties of the N-terminal DNA binding domain within the large subunit of replication factor C.

Replication Factor C (RFC) is a five-subunit protein complex required for eukaryotic DNA replication and repair. The large subunit within this complex contains a C-terminal DNA binding domain which provides specificity for PCNA loading at a primer-template and a second, N-terminal DNA binding domain of unknown function. We isolated the N-terminal DNA binding domain from Drosophila melanogaster and defined the region within this polypeptide required for DNA binding. The DNA determinants most efficiently recognized by both the Drosophila minimal DNA binding domain and the N-terminal half of the human large subunit consist of a double-stranded DNA containing a recessed 5' phosphate. DNA containing a recessed 5' phosphate was preferred 5-fold over hairpined DNA containing a recessed 3' hydroxyl. Combined with existing data, these DNA binding properties suggest a role for the N-terminal DNA binding domain in the recognition of phosphorylated DNA ends.     

Repercussions of the COVID-19 pandemic on athletes: a cross-sectional study

The COVID-19 pandemic has presented significant challenges and implications for the sports community. Thus, this study aimed to describe the prevalence of COVID-19 in Brazilian athletes and identify the epidemiological, clinical, athletic, life and health factors associated with the disease in these individuals. A cross-sectional study was performed involving 414 athletes from 22 different sports using an online questionnaire from August to November 2020. The association between the athletes’ characteristics and COVID-19 was evaluated using a logistic regression model. The prevalence of COVID-19 was 8.5%, although only 40% of athletes reported having been tested. Being under 27 years of age (3-fold), having children (~5-fold), having a teammate test positive for COVID-19 (2.5-fold), and smoking (14-fold) were associated with a possible higher risk of disease. Almost 20% of athletes self-reported musculoskeletal injuries during the period of the pandemic that was studied. Athletes with a university education (P = 0.02), a profession other than sports (P < 0.001), those from a low-income family (P = 0.01), and public health system users (P = 0.04) were significantly less frequently tested for COVID-19, whereas international competitors, athletes who received a wage, and athletes who had a teammate who tested positive for COVID-19 were 2-, 3-, and 15-fold more likely to be tested for COVID-19, respectively. Approximately 26% of the athletes who tested negative or were untested reported more than three characteristic COVID-19 symptoms, and 11% of athletes who tested positive for COVID-19 were asymptomatic. The identification of modifiable (have children, smoking, and teammates positively tested) and non-modifiable (age under 27 years) factors related to COVID-19 in athletes can contribute to implementing surveillance programmes to decrease the incidence of COVID-19 in athletes and its negative impacts in sports.     

COMP mutations: domain-dependent relationship between abnormal chondrocyte trafficking and clinical PSACH and MED phenotypes

Mutations in cartilage oligomeric matrix protein (COMP) produce clinical phenotypes ranging from the severe end of the spectrum, pseudoachondroplasia (PSACH), which is a dwarfing condition, to a mild condition, multiple epiphyseal dysplasia (MED). Patient chondrocytes have a unique morphology characterized by distended rER cisternae containing lamellar deposits of COMP and other extracellular matrix proteins. It has been difficult to determine why different mutations give rise to variable clinical phenotypes. Using our in vitro cell system, we previously demonstrated that the most common PSACH mutation, D469del, severely impedes trafficking of COMP and type IX collagen in chondrocytic cells, consistent with observations from patient cells. Here, we hypothesize that PSACH and MED mutations variably affect the cellular trafficking behavior of COMP and that the extent of defective trafficking correlates with clinical phenotype. Twelve different recombinant COMP mutations were expressed in rat chondrosarcoma cells and the percent cells with ER-retained COMP was assessed. For mutations in type 3 (T3) repeats, trafficking defects correlated with clinical phenotype; PSACH mutations had more cells retaining mutant COMP, while MED mutations had fewer. In contrast, the cellular trafficking pattern observed for mutations in the C-terminal globular domain (CTD) was not predictive of clinical phenotype. The results demonstrate that different COMP mutations in the T3 repeat domain have variable effects on intracellular transport, which correlate with clinical severity, while CTD mutations do not show such a correlation. These findings suggest that other unidentified factors contribute to the effect of the CTD mutations. J. Cell. Biochem. 103: 778-787, 2008. (c) 2007 Wiley-Liss, Inc.     

Endocytic recycling proteins EHD1 and EHD2 interact with fer-1-like-5 (Fer1L5) and mediate myoblast fusion

The mammalian ferlins are calcium-sensing, C2 domain-containing proteins involved in vesicle trafficking. Myoferlin and dysferlin regulate myoblast fusion and muscle membrane resealing, respectively. Correspondingly, myoferlin is most highly expressed in singly nucleated myoblasts, whereas dysferlin expression is increased in mature, multinucleated myotubes. Myoferlin also mediates endocytic recycling and participates in trafficking the insulin-like growth factor receptor. We have now characterized a novel member of the ferlin family, Fer1L5, because of its high homology to dysferlin and myoferlin. We found that Fer1L5 protein is expressed in small myotubes that contain only two to four nuclei. We also found that Fer1L5 protein binds directly to the endocytic recycling proteins EHD1 and EHD2 and that the second C2 domain in Fer1L5 mediates this interaction. Reduction of EHD1 and/or EHD2 inhibits myoblast fusion, and EHD2 is required for normal translocation of Fer1L5 to the plasma membrane. The characterization of Fer1L5 and its interaction with EHD1 and EHD2 underscores the complex requirement of ferlin proteins and mediators of endocytic recycling for membrane trafficking events during myotube formation.     

Sediment quality of North Carolina estuaries: an integrative assessment of sediment contamination, toxicity, and condition of benthic fauna

Sediment quality of North Carolina estuaries was evaluated using synoptic data on sediment chemistry, toxicity, and macroinfaunal community structure from 175 subtidal stations sampled during the summers of 1994–1997. The study area included Currituck, Albemarle, and Pamlico Sounds; estuarine portions of major rivers (e.g., Chowan, Roanoke, Tar-Pamlico, Neuse, New, Cape Fear); and numerous smaller tributaries and coastal embayments between the Virginia and South Carolina borders. A probabilistic sampling design permitted statistical estimation of the spatial extent of degraded versus non-degraded condition across these estuaries. Over half (54 ± 7%) of the surveyed area had high sediment quality characterized by healthy benthic assemblages and low levels of sediment contamination and toxicity. The remaining 46% showed evidence of significant stress in one or more of the above sediment-quality-triad components. While this is a sizable area, portions of it (27 ± 6%) were represented by sites with no connection between presence of stressors and adverse biological responses. Only 19% of the total area showed evidence of an impaired benthos coupled to significant pollution exposure (high sediment contamination, toxicity, or both). Impaired benthic condition was more closely linked to sediment contamination than to low dissolved oxygen (based on instantaneous oxygen measurements). The most pervasive contaminants were the metals arsenic, mercury, chromium, and nickel; the pesticides lindane, dieldrin, DDT, and DDT derivatives; and total PCBs. Degraded condition in all three components of the sediment quality triad co-occurred in <10% of the study area, suggesting that strong contaminant-induced effects on the benthos are limited to a small (yet ecologically significant) percentage of total estuarine area. The spatial extent of sediment contamination and toxicity was much less in these estuaries in comparison to other U.S. coastal regions where similar studies have been performed.     

Collaborative research and intellectual property rights

Scientists find themselves working more and more with indigenous, traditional and local communities in all aspects of their collections and investigations. Indigenous knowledge has become increasingly important in research while at the same time local communities have become increasingly politicized in the use, misappropriation, and commercialisation of their knowledge and biogenetic resources. It is becoming more and more difficult for even the most well-intentioned scientists to stride into indigenous areas and collect plants, animals, folk tales, and photos without having first to convince local leaders that the scholarly efforts will somehow benefit the communities — that the benefits of research results will directly and indirectly lead to strengthening the traditional society. In many parts of the world, indigenous peoples only allow Collaborative Research in which the scientific priorities and agendas are controlled by the communities, or Community Controlled Research in which the communities actually contract scientists to carry out the group's research plan. Control over data has become one of the key battle cries for the indigenous movement, that is now demanding Intellectual Property Rights over information obtained through research and just compensation for economic benefits that eventually may accrue. This paper deals with some of the ethical and practical issues that frame this rapidly evolving debate.     

A histological and immunohistochemical study of the effects of N-acetyl cysteine on retinopathy of prematurity by modifying insulin-like growth factor-1

Retinopathy of prematurity (ROP) is a vasoproliferative disorder that occurs in premature infants and may lead to permanent visual impairment. We investigated both the possible protective role of N-acetyl cysteine (NAC) for preventing ROP and the role of IGF-1 in the disorder. Forty-five newborn rats were divided into three groups. Group 1 was raised in room air as controls. Group 2 was exposed to 60% oxygen for 14 days after birth, then transferred to room air. Group 3 was exposed to the same conditions as group 2, but received intraperitoneal injections of NAC on postnatal days 7–17. After 35 days, both eyes of all rats were processed for histology. Some sections were stained with hematoxylin and eosin to assess structural changes and other sections were immunostained to determine the location of IGF-1. Frozen sections also were prepared and stained for adenosine triphosphatase to detect retinal blood vessels. Compared to the controls, more blood vessels, many of which were abnormal, and increased IGF-1 expression were observed in group 2. In group 3, abnormal blood vessels and IGF-1 expression were less evident. NAC appeared to be an effective vascular-protective agent for ROP by decreasing IGF-1 expression.