Confirmatory Factor Analysis of the Spirituality and Spiritual Care Rating Scale: A Cross-Sectional Study in Eight US Nursing Homes.

The aim of this study was to test a modified Spirituality and Spiritual Care Rating Scale (SSCRS) and report initial findings, based on a cross-sectional descriptive survey from eight nursing homes in the USA. This study examined the psychometric properties of a modified version of the SSCRS for assessing the perceptions of staff (i.e. nurses, rehabilitation staff, food and nutrition service staff, activities staff, social workers, and administrative staff) who work in nursing homes. The modified version of the SSCRS is measuring respondents' perceptions of spirituality, spiritual care, religiosity, and personalized care. Confirmatory factor analysis showed satisfactory goodness of fit for the original four-factor structure of the SSCRS. The modified version demonstrated good internal consistency and reliability (Cronbach's alpha ranging from 0.71 to 0.90). Initial findings showed statistically significant differences across all four sub-scales. Further research is needed to psychometrically test the modified tool.

MRB3010 is a core component of the MRB1 complex that facilitates an early step of the kinetoplastid RNA editing process.

Gene expression in the mitochondria of the kinetoplastid parasite Trypanosoma brucei is regulated primarily post-transcriptionally at the stages of RNA processing, editing, and turnover. The mitochondrial RNA-binding complex 1 (MRB1) is a recently identified multiprotein complex containing components with distinct functions during different aspects of RNA metabolism, such as guide RNA (gRNA) and mRNA turnover, precursor transcript processing, and RNA editing. In this study we examined the function of the MRB1 protein, Tb927.5.3010, which we term MRB3010. We show that MRB3010 is essential for growth of both procyclic form and bloodstream form life-cycle stages of T. brucei. Down-regulation of MRB3010 by RNAi leads to a dramatic inhibition of RNA editing, yet its depletion does not impact total gRNA levels. Rather, it appears to affect the editing process at an early stage, as indicated by the accumulation of pre-edited and small partially edited RNAs. MRB3010 is present in large (>20S) complexes and exhibits both RNA-dependent and RNA-independent interactions with other MRB1 complex proteins. Comparison of proteins isolated with MRB3010 tagged at its endogenous locus to those reported from other MRB1 complex purifications strongly suggests the presence of an MRB1 "core" complex containing five to six proteins, including MRB3010. Together, these data further our understanding of the function and composition of the imprecisely defined MRB1 complex.

A novel member of the RNase D exoribonuclease family functions in mitochondrial guide RNA metabolism in Trypanosoma brucei.

RNA turnover and RNA editing are essential for regulation of mitochondrial gene expression in Trypanosoma brucei. RNA turnover is controlled in part by RNA 3' adenylation and uridylation status, with trans-acting factors also impacting RNA homeostasis. However, little is known about the mitochondrial degradation machinery or its regulation in T. brucei. We have identified a mitochondrial exoribonuclease, TbRND, whose expression is highly up-regulated in the insect proliferative stage of the parasite. TbRND shares sequence similarity with RNase D family enzymes but differs from all reported members of this family in possessing a CCHC zinc finger domain. In vitro, TbRND exhibits 3' to 5' exoribonuclease activity, with specificity toward uridine homopolymers, including the 3' oligo(U) tails of guide RNAs (gRNAs) that provide the sequence information for RNA editing. Several lines of evidence generated from RNAi-mediated knockdown and overexpression cell lines indicate that TbRND functions in gRNA metabolism in vivo. First, TbRND depletion results in gRNA tails extended by 2-3 nucleotides on average. Second, overexpression of wild type but not catalytically inactive TbRND results in a substantial decrease in the total gRNA population and a consequent inhibition of RNA editing. The observed effects on the gRNA population are specific as rRNAs, which are also 3'-uridylated, are unaffected by TbRND depletion or overexpression. Finally, we show that gRNA binding proteins co-purify with TbRND. In summary, TbRND is a novel 3' to 5' exoribonuclease that appears to have evolved a function highly specific to the mitochondrion of trypanosomes.

Structure of the Trypanosoma brucei p22 protein, a cytochrome oxidase subunit II-specific RNA-editing accessory factor.

Kinetoplastid RNA (k-RNA) editing is a complex process in the mitochondria of kinetoplastid protozoa, including Trypanosoma brucei, that involves the guide RNA-directed insertion and deletion of uridines from precursor-mRNAs to produce mature, translatable mRNAs. k-RNA editing is performed by multiprotein complexes called editosomes. Additional non-editosome components termed k-RNA-editing accessory factors affect the extent of editing of specific RNAs or classes of RNAs. The T. brucei p22 protein was identified as one such accessory factor. Here we show that p22 contributes to cell growth in the procyclic form of T. brucei and functions as a cytochrome oxidase subunit II-specific k-RNA-editing accessory factor. To gain insight into its functions, we solved the crystal structure of the T. brucei p22 protein to 2.0-A resolution. The p22 structure consists of a six-stranded, antiparallel beta-sheet flanked by five alpha-helices. Three p22 subunits combine to form a tight trimer that is primarily stabilized by interactions between helical residues. One side of the trimer is strikingly acidic, while the opposite face is more neutral. Database searches show p22 is structurally similar to human p32, which has a number of functions, including regulation of RNA splicing. p32 interacts with a number of target proteins via its alpha1 N-terminal helix, which is among the most conserved regions between p22 and p32. Co-immunoprecipitation studies showed that p22 interacts with the editosome and the k-RNA accessory protein, TbRGG2, and alpha1 of p22 was shown to be important for the p22-TbRGG2 interaction. Thus, these combined studies suggest that p22 mediates its role in k-RNA editing by acting as an adaptor protein.

Differential contribution of TAP and tapasin to HLA class I antigen expression.

Expression of class I human leucocyte antigens (HLA) on the surface of malignant cells is critical for their recognition and destruction by cytotoxic T lymphocytes. Surface expression requires assembly and folding of HLA class I molecules in the endoplasmic reticulum with the assistance of proteins such as Transporter associated with Antigen Processing (TAP) and tapasin. Interferon-gamma induces both TAP and tapasin so dissection of which protein contributes more to HLA class I expression has not been possible previously. In this study, we take advantage of a human melanoma cell line in which TAP can be induced, but tapasin cannot. Interferon-gamma increases TAP protein levels dramatically but HLA class I expression at the cell surface does not increase substantially, indicating that a large increase in peptide supply is not sufficient to increase HLA class I expression. On the other hand, transfection of either allelic form of tapasin (R240 or T240) enhances HLA-B*5001 and HLA-B*5701 antigen expression considerably with only a modest increase in TAP. Together, these data indicate that in the presence of minimal TAP activity, tapasin can promote substantial HLA class I expression at the cell surface.

Proteasomes and proteasome activator 200 kDa (PA200) accumulate on chromatin in response to ionizing radiation.

Proteasome activator 200 kDa (PA200) forms nuclear foci after exposure of cells to ionizing radiation and enhances proteasome activity in vitro. Within cells, it is unclear whether PA200 responds to radiation alone or in association with proteasomes. In the present study, we identified three forms of cellular PA200 (termed PA200i, ii and iii) at the mRNA and protein levels. Neither PA200ii nor PA200iii appears to associate with proteasomes. All detectable PA200i is associated with proteasomes, which indicates that PA200i and proteasomes function together within the cell. Consistent with this idea, we find that exposure of cells to radiation leads to an equivalent accumulation of both PA200i and core proteasomes on chromatin. This increase in PA200 and proteasomes on chromatin is not specific to the stage of cell cycle arrest since it occurs in cells that arrest in G(2)/M and cells that arrest in G(1)/S after exposure to radiation. These data provide evidence that PA200 and proteasomes function together within cells and respond to a specific radiation-induced damage independent of the stage of cell cycle arrest.

Patient Perspectives on Participation in Cognitive Functional Therapy for Chronic Low Back Pain.

BACKGROUND: Cognitive functional therapy (CFT) has been shown to reduce pain and disability in people with chronic low back pain. OBJECTIVES: The purpose of this study was to investigate participants' experience of CFT by comparing participants who reported differing levels of improvement after participation in CFT, potentially yielding insight into the implementation of this approach. DESIGN: This was a noninterventional, cross-sectional, qualitative study with an interpretive description framework. METHODS: Individuals who had participated in CFT in 2 physical therapy settings (in Ireland and Australia) were recruited through purposive sampling based on disability outcomes postintervention (n=9), and theoretical sampling (n=5). This sampling strategy was used to capture a range of participant experiences but was not used to define the final qualitative groupings. Semistructured interviews were conducted 3 to 6 months postintervention. RESULTS: Three groups emerged from the qualitative analysis: large improvers, small improvers, and unchanged. Two themes encapsulating the key requirements in achieving a successful outcome through CFT were identified: (1) changing pain beliefs and (2) achieving independence. Changing pain beliefs to a more biopsychosocial perspective required a strong therapeutic alliance, development of body awareness, and the experience of control over pain. Independence was achieved by large improvers through newly cultivated problem-solving skills, self-efficacy, decreased fear of pain, and improved stress coping. Residual fear and poor stress coping meant that small improvers were easily distressed and lacked independence. Those who were unchanged continued to feel defined by their pain and retained a biomedical perspective. CONCLUSIONS: A successful outcome after CFT is dependent on instilling biopsychosocial pain beliefs and developing independence among participants. Small improvers may require ongoing support to maintain results. Further study is needed to elucidate the optimal approach for those who were unchanged.

The signature of fine scale local adaptation in Atlantic salmon revealed from common garden experiments in nature.

Understanding the extent, scale and genetic basis of local adaptation (LA) is important for conservation and management. Its relevance in salmonids at microgeographic scales, where dispersal (and hence potential gene flow) can be substantial, has however been questioned. Here, we compare the fitness of communally reared offspring of local and foreign Atlantic salmon Salmo salar from adjacent Irish rivers and reciprocal F1 hybrid crosses between them, in the wild 'home' environment of the local population. Experimental groups did not differ in wild smolt output but a catastrophic flood event may have limited our ability to detect freshwater performance differences, which were evident in a previous study. Foreign parr exhibited higher, and hybrids intermediate, emigration rates from the natal stream relative to local parr, consistent with genetically based behavioural differences. Adult return rates were lower for the foreign compared to the local group. Overall lifetime success of foreigners and hybrids relative to locals was estimated at 31% and 40% (mean of both hybrid groups), respectively. The results imply a genetic basis to fitness differences among populations separated by only 50 km, driven largely by variation in smolt to adult return rates. Hence even if supplementary stocking programs obtain broodstock from neighbouring rivers, the risk of extrinsic outbreeding depression may be high.

Additive and transcript-specific effects of KPAP1 and TbRND activities on 3' non-encoded tail characteristics and mRNA stability in Trypanosoma brucei.

Short, non-encoded oligo(A), oligo(U), or A/U tails can impact mRNA stability in kinetoplastid mitochondria. However, a comprehensive picture of the relative effects of these modifications in RNA stability is lacking. Furthermore, while the U-preferring exoribonuclease TbRND acts on U-tailed gRNAs, its role in decay of uridylated mRNAs has only been cursorily investigated. Here, we analyzed the roles of mRNA 3' tail composition and TbRND in RNA decay using cells harbouring single or double knockdown of TbRND and the KPAP1 poly(A) polymerase. Analysis of mRNA abundance and tail composition reveals dramatic and transcript-specific effects of adenylation and uridylation on mitochondrial RNAs. Oligo(A) and A-rich tails can stabilize a proportion of edited and never-edited RNAs. However, non-tailed RNAs are not inherently unstable, implicating additional stability determinants and/or spatial segregation of sub-populations of a given RNA in regulation of RNA decay. Oligo(U) tails, which have been shown to contribute to decay of some never-edited RNAs, are not universally destabilizing. We also show that RNAs display very different susceptibility to uridylation in the absence of KPAP1, a factor that may contribute to regulation of decay. Finally, 3' tail composition apparently impacts the ability of an RNA to be edited.

What is the role of alternate splicing in antigen presentation by major histocompatibility complex class I molecules?

The expression of major histocompatibility complex (MHC) class I molecules on the cell surface is critical for recognition by cytotoxic T lymphocytes (CTL). This recognition event leads to destruction of cells displaying MHC class I-viral peptide complexes or cells displaying MHC class I-mutant peptide complexes. Before they can be transported to the cell surface, MHC class I molecules must associate with their peptide ligand in the endoplasmic reticulum (ER) of the cell. Within the ER, numerous proteins assist in the appropriate assembly and folding of MHC class I molecules. These include the heterodimeric transporter associated with antigen processing (TAP1 and TAP2), the heterodimeric chaperone-oxidoreductase complex of tapasin and ERp57 and the general ER chaperones calreticulin and calnexin. Each of these accessory proteins has a well-defined role in antigen presentation by MHC class I molecules. However, alternate splice forms of MHC class I heavy chains, TAP and tapasin, have been reported suggesting additional complexity to the picture of antigen presentation. Here, we review the importance of these different accessory proteins and the progress in our understanding of alternate splicing in antigen presentation.

Identification of an alternate splice form of tapasin in human melanoma.

Assembly of major histocompatibility complex (MHC) class I molecules with peptide in the endoplasmic reticulum requires the assistance of tapasin. Alternative splicing, which is known to regulate many genes, has been reported for tapasin only in the context of mutations. Here, we report on an alternate splice form of tapasin (tpsnΔEx3) derived from a human melanoma cell line that does not appear to be caused by mutations. Excision of exon 3 results in deletion of amino acids 70 to 156 within the beta barrel region, but the membrane proximal Ig domain, the transmembrane domain, and cytoplasmic tail of tapasin are intact. Introduction of tpsnΔEx3 into a tapasin-deficient cell line does not restore MHC class I expression at the cell surface. Similar to a previously described tapasin mutant (tpsnΔN50), tpsnΔEx3 interacts with TAP. Therefore, we used these altered forms of tapasin to test the importance of MHC class I interaction with TAP. In the presence of wild-type tapasin, transfection of tpsnΔN50, but not tpsnΔEx3, reduced MHC class I expression at the cell surface likely due its ability to compete MHC class I molecules from TAP. Together these findings suggest that tumor cells may contain alternate splice forms of tapasin which may regulate MHC class I antigen presentation.