Quantitative phosphoproteomics reveals the role of protein arginine phosphorylation in the bacterial stress response.

Arginine phosphorylation is an emerging protein modification implicated in the general stress response of Gram-positive bacteria. The modification is mediated by the arginine kinase McsB, which phosphorylates and inactivates the heat shock repressor CtsR. In this study, we developed a mass spectrometric approach accounting for the peculiar chemical properties of phosphoarginine. The improved methodology was used to analyze the dynamic changes in the Bacillus subtilis arginine phosphoproteome in response to different stress situations. Quantitative analysis showed that a B. subtilis mutant lacking the YwlE arginine phosphatase accumulated a strikingly large number of arginine phosphorylations (217 sites in 134 proteins), however only a minor fraction of these sites was increasingly modified during heat shock or oxidative stress. The main targets of McsB-mediated arginine phosphorylation comprise central factors of the stress response system including the CtsR and HrcA heat shock repressors, as well as major components of the protein quality control system such as the ClpCP protease and the GroEL chaperonine. These findings highlight the impact of arginine phosphorylation in orchestrating the bacterial stress response.

Structural basis for recognizing phosphoarginine and evolving residue-specific protein phosphatases in gram-positive bacteria.

Many cellular pathways are regulated by the competing activity of protein kinases and phosphatases. The recent identification of arginine phosphorylation as a protein modification in bacteria prompted us to analyze the molecular basis of targeting phospho-arginine. In this work, we characterize an annotated tyrosine phosphatase, YwlE, that counteracts the protein arginine kinase McsB. Strikingly, structural studies of YwlE reaction intermediates provide a direct view on a captured arginine residue. Together with biochemical data, the crystal structures depict the evolution of a highly specific phospho-arginine phosphatase, with the use of a size-and-polarity filter for distinguishing phosphorylated arginine from other phosphorylated side chains. To confirm the proposed mechanism, we performed bioinformatic searches for phosphatases, employing a similar selectivity filter, and identified a protein in Drosophila melanogaster exhibiting robust arginine phosphatase activity. In sum, our findings uncover the molecular framework for specific targeting of phospho-arginine and suggest that protein arginine (de)phosphorylation may be relevant in eukaryotes.

Infant oral health care: beliefs and practices of American Academy of Pediatric Dentistry members.

PURPOSE: The purpose of this study was to survey American Academy of Pediatric Dentistry (AAPD) members about their beliefs and practices regarding the age 1 dental visit. METHODS: A 31-item survey was mailed to 1,500 randomly selected AAPD members to assess their beliefs and practices regarding the age 1 dental visit and prenatal dental counseling. Descriptive analyses and chi-square tests were done to analyze the data. RESULTS: Five hundred fifty-four (~37%) surveys were returned; 496 (91%) respondents agreed with the AAPD Policy on the Dental Home, while 490 (~90%) respondents performed age 1 dental evaluations. The most commonly cited reason respondents gave for not performing infant evaluations was "parents do not see the value." Eighty-five (~15 %) respondents provided prenatal dental counseling to members in their community, and 250 (~51%) who performed infant oral evaluations inquired about parents' dental history. Younger, more recently trained pediatric dentists were more likely to agree with the policy compared to older, more experienced pediatric dentists. CONCLUSIONS: Most respondents agreed with the American Academy of Pediatric Dentistry's Policy on the Dental Home and performed infant oral evaluations. Few respondents (15%) provided prenatal dental counseling to individuals/groups in their community.

Survey of pediatric dentists' own oral health behavior.

Oral health behavior is voluntary and, therefore, requires motivation. Dentists serve as role models for their patients. We sought to determine the pediatric dentists' own adherence to oral health behavior recommendations through a 14-question survey administered to 129 pediatric dentists. A significant percentage of respondents fell short in brushing, flossing and regularity of dental exams. A trend was seen for decreasing exam frequency with decreasing population density and for increasing brushing frequency with increasing age. Although most dentists follow their profession's oral health guidelines, a significant percentage does not.

McsB is a protein arginine kinase that phosphorylates and inhibits the heat-shock regulator CtsR.

All living organisms face a variety of environmental stresses that cause the misfolding and aggregation of proteins. To eliminate damaged proteins, cells developed highly efficient stress response and protein quality control systems. We performed a biochemical and structural analysis of the bacterial CtsR/McsB stress response. The crystal structure of the CtsR repressor, in complex with DNA, pinpointed key residues important for high-affinity binding to the promoter regions of heat-shock genes. Moreover, biochemical characterization of McsB revealed that McsB specifically phosphorylates arginine residues in the DNA binding domain of CtsR, thereby impairing its function as a repressor of stress response genes. Identification of the CtsR/McsB arginine phospho-switch expands the repertoire of possible protein modifications involved in prokaryotic and eukaryotic transcriptional regulation.

Comparison of nutritional counseling between provider types.

OBJECTIVE: The goal of this study was to compare the provision of and attitudes toward nutritional counseling between pediatricians and pediatric dentists. METHODS: Questionnaires were mailed to 500 US members of the American Academy of Pediatric Dentistry and the American Academy of Pediatrics. RESULTS: The survey had a response rate of 33% (pediatric dentists=36%; pediatricians=29%). 84% of pediatricians routinely offered nutritional counseling compared to 71% of pediatric dentists. With respect to cariogenic foods, 82% of pediatric dentists and 74% of pediatricians correctly recommended bottle contents, and 33% of pediatric dentists and 19% of pediatricians correctly recommended snacks. 79% of pediatric dentists compared to 69% of pediatricians believed nutritional counseling had at least some effectiveness. Eighty-nine percent of both specialties somewhat or strongly agreed that nutritional counseling was an important aspect of oral health care. CONCLUSIONS: While pediatric dentists made slightly better recommendations than pediatricians regarding cariogenic foods, both provider types demonstrated a need for more education regarding nutritional counseling.

Attitudes of Chinese parents toward the oral health of their children with caries: a qualitative study.

PURPOSE: The purpose of this study was to investigate Chinese parents' beliefs and perspectives regarding extensive caries (EC), oral care habits, and dental treatment. An overview of qualitative research methods is provided. METHODS: In this qualitative study, 20 in-depth interviews were conducted with parents of children diagnosed with EC. Parents lived in a major metropolitan area, and many were newly immigrated. Parents who accepted or refused dental treatment for their children under general anesthesia or sedation were included. Transcribed interviews were analyzed using standard thematic analysis. RESULTS: Negative themes were: (1) fear of dental anesthesia and its adverse effects; (2) parents' own lack of dental education as children; (3) lack of social support in seeking dental treatment; (4) inadequate knowledge of oral hygiene; and (5) cultural beliefs that did not support the preservation of the primary dentition. Positive themes were: (1) trust in the providers and in Western medicine; and (2) satisfaction with outcomes of dental treatment. CONCLUSIONS: Several factors were found that could contribute to a higher rate of EC in this population. Providers can benefit from this study by anticipating what practices and attitudes are common in this community. Earlier intervention and delivery of culturally sensitive care can prevent or delay progression of this dental disease.