Protein phosphatase 1 (PP1) is expressed in all eukaryotic cells and catalyzes a substantial fraction of phosphoserine/threonine dephosphorylation reactions. It forms stable complexes with PP1-interacting proteins (PIPs) that guide the phosphatase throughout its life cycle and control its fate and function. The diversity of PIPs is huge (≈200 in vertebrates), and most of them combine short linear motifs to form large and unique interaction interfaces with PP1. Many PIPs have separate domains for PP1 anchoring, PP1 regulation, substrate recruitment and subcellular targeting, which enable them to direct associated PP1 to a specific subset of substrates and mediate acute activity control. Hence, PP1 functions as the catalytic subunit of a large number of multimeric holoenzymes, each with its own subset of substrates and mechanism(s) of regulation.

Whereas telephone interventions only increased spontaneous reporting in the first 4 months of follow-up, workshops significantly increased both the quantity and relevance of spontaneous ADR reporting for more than 1 year.

Objectives Antibiotic misprescription is a major driver of resistance, which is a worldwide public health problem. Therefore, our aim is to assess the influence of the determinants of physician prescribing on the quality of antibiotic use. Methods A 3 year cohort study including all primary-care physicians working in Portugal's Central Regional Health Administration (n = 1094) was conducted. We assessed the determinants of prescribing using a pre-validated, personally addressed, reply-paid, self-administered questionnaire (sent four times to non-responders, between September 2011 and February 2012) designed to collect information on physicians' attitudes to and knowledge of antibiotic prescribing as well as their socio-demographic and professional data. To evaluate antibiotic prescribing, we've calculated ESAC 12 quality indicators per physician per year, allowing us to stratify them as good or poor prescribers according to their performance on those indicators. Associations between determinants and outcomes were fitted with generalized linear mixed models. Results The overall response rate was 46.1%. Emergency activity (OR [95% CI] = 0.29 [0.16-0.54]; p < 0.05) and workload (number of patients seen per day: OR [95% CI] = 0.97 [0.94-1.00]; p < 0.05; number of patients seen per week in emergencies: OR [95% CI] = 0.98 [0.97-0.99]; p < 0.05) were both related to poor quality of antibiotic prescribing. Statistically significant odds ratios were also obtained for ignorance (IqOR [95% CI] = 2.14 [1.31-3.52]), complacency (1/IqOR [95% CI] = 1.19 [1.01-1.41]) and responsibility of others (1/IqOR [95% CI] = 1.78 [1.10-3.06]). Conclusions The above results serve to emphasize workload, working at emergency departments and physicians' attitudes identified as critical factors affecting antibiotic prescribing. This provides new insights for clinicians, researchers and policy makers when it comes to developing and improving the clinical and economic outcomes of antibiotic use. Key limitations of the study included the difficulty of results extrapolation and the limitations of the stratification method based on the antibiotic prescribing quality indicators.

BackgroundUnderstanding physicians’ antibiotic-prescribing behaviour is fundamental when it comes to improving antibiotic use and tackling the growing rates of antimicrobial resistance. The aim of the study was to develop and validate -in terms of face validity, content validity and reliability- an instrument designed to assess the attitudes and knowledge underlying physician antibiotic prescribing.MethodsThe questionnaire development and validation process comprised two different steps, namely: (1) content and face validation, which included a literature review and validation both by physicians and by Portuguese language and clinical psychology experts; and (2) reliability analysis, using the test-retest method, to assess the questionnaire’s internal consistency (Cronbach’s alpha) and reproducibility (intraclass correlation coefficient - ICC). The questionnaire includes 17 items assessing attitudes and knowledge about antibiotic prescribing and resistances and 9 items evaluating the importance of different sources of knowledge. The study was conducted in the catchment area covered by Portugal’s Northern Regional Health Administration and used a convenience sample of 61 primary-care and 50 hospital-care physicians.ResultsResponse rate was 64 % (49 % to retest) for primary-care physicians and 66 % (60 % to retest) for hospital-care physicians. Content validity resulted in 9 changes to professional concepts. Face validity assessment resulted in 19 changes to linguistic and interpretative terms. In the case of the reliability analysis, the ICC values indicated a minimum of fair to good reproducibility (ICC > 0.4), and the Cronbach alpha values were satisfactory (α > 0.70).ConclusionsThe questionnaire developed is valid -in terms of face validity, content validity and reliability- for assessing physicians’ attitudes to and knowledge of antibiotic prescribing and resistance, in both hospital and primary-care settings, and could be a very useful tool for characterising physicians’ antibiotic-prescribing behaviour.Electronic supplementary materialThe online version of this article (doi:10.1186/s12879-015-1332-y) contains supplementary material, which is available to authorized users.

NIPP1 is one of the major nuclear interactors of protein phosphatase PP1. The deletion of NIPP1 in mice is early embryonic lethal, which has precluded functional studies in adult tissues. Hence, we have generated an inducible NIPP1 knockout model using a tamoxifen-inducible Cre recombinase transgene. The inactivation of the NIPP1 encoding alleles (Ppp1r8) in adult mice occurred very efficiently in testis and resulted in a gradual loss of germ cells, culminating in a Sertoli-cell only phenotype. Before the overt development of this phenotype Ppp1r8 −/− testis showed a decreased proliferation and survival capacity of cells of the spermatogenic lineage. A reduced proliferation was also detected after the tamoxifen-induced removal of NIPP1 from cultured testis slices and isolated germ cells enriched for undifferentiated spermatogonia, hinting at a testis-intrinsic defect. Consistent with the observed phenotype, RNA sequencing identified changes in the transcript levels of cell-cycle and apoptosis regulating genes in NIPP1-depleted testis. We conclude that NIPP1 is essential for mammalian spermatogenesis because it is indispensable for the proliferation and survival of progenitor germ cells, including (un)differentiated spermatogonia.

Protein phosphatase 1 (PP1) catalyzes more than half of all phosphoserine/threonine dephosphorylation reactions in mammalian cells. In vivo PP1 does not exist as a free catalytic subunit but is always associated with at least one regulatory PP1-interacting protein (PIP) to generate a large set of distinct holoenzymes. Each PP1 complex controls the dephosphorylation of only a small subset of PP1 substrates. We screened the literature for genetically engineered mouse models and identified models for all PP1 isoforms and 104 PIPs. PP1 itself and at least 49 PIPs were connected to human disease-associated phenotypes. Additionally, phenotypes related to 17 PIPs were clearly linked to altered PP1 function, while such information was lacking for 32 other PIPs. We propose structural reverse genetics, which combines structural characterization of proteins with mouse genetics, to identify new PP1-related therapeutic targets. The available mouse models confirm the pleiotropic action of PP1 in health and diseases.

Highlights d The LRRs of SDS22 interact with PP1 helices a5 and a6 d A basic surface patch of SDS22 binds phosphorylated BCLAF1 d Many PP1 holoenzymes contain SDS22 as a third subunit d SDS22 prevents the autodephosphorylation of PP1

BackgroundProtein Ser/Thr Phosphatase PPP1CC2 is an alternatively spliced isoform of PPP1C that is highly enriched in testis and selectively expressed in sperm. Addition of the phosphatase inhibitor toxins okadaic acid or calyculin A to caput and caudal sperm triggers and stimulates motility, respectively. Thus, the endogenous mechanisms of phosphatase inhibition are fundamental for controlling sperm function and should be characterized. Preliminary results have shown a protein phosphatase inhibitor activity resembling PPP1R2 in bovine and primate spermatozoa.ResultsHere we show conclusively, for the first time, that PPP1R2 is present in sperm. In addition, we have also identified a novel protein, PPP1R2P3. The latter was previously thought to be an intron-less pseudogene. We show that the protein corresponding to the pseudogene is expressed. It has PPP1 inhibitory potency similar to PPP1R2. The potential phosphosites in PPP1R2 are substituted by non-phosphorylable residues, T73P and S87R, in PPP1R2P3. We also confirm that PPP1R2/PPP1R2P3 are phosphorylated at Ser121 and Ser122, and report a novel phosphorylation site, Ser127. Subfractionation of sperm structures show that PPP1CC2, PPP1R2/PPP1R2P3 are located in the head and tail structures.ConclusionsThe conclusive identification and localization of sperm PPP1R2 and PPP1R2P3 lays the basis for future studies on their roles in acrosome reaction, sperm motility and hyperactivation. An intriguing possibility is that a switch in PPP1CC2 inhibitory subunits could be the trigger for sperm motility in the epididymis and/or sperm hyperactivation in the female reproductive tract.