The complete sequence and comparative analysis of ape sex chromosomes.

Apes possess two sex chromosomes-the male-specific Y chromosome and the X chromosome, which is present in both males and females. The Y chromosome is crucial for male reproduction, with deletions being linked to infertility1. The X chromosome is vital for reproduction and cognition2. Variation in mating patterns and brain function among apes suggests corresponding differences in their sex chromosomes. However, owing to their repetitive nature and incomplete reference assemblies, ape sex chromosomes have been challenging to study. Here, using the methodology developed for the telomere-to-telomere (T2T) human genome, we produced gapless assemblies of the X and Y chromosomes for five great apes (bonobo (Pan paniscus), chimpanzee (Pan troglodytes), western lowland gorilla (Gorilla gorilla gorilla), Bornean orangutan (Pongo pygmaeus) and Sumatran orangutan (Pongo abelii)) and a lesser ape (the siamang gibbon (Symphalangus syndactylus)), and untangled the intricacies of their evolution. Compared with the X chromosomes, the ape Y chromosomes vary greatly in size and have low alignability and high levels of structural rearrangements-owing to the accumulation of lineage-specific ampliconic regions, palindromes, transposable elements and satellites. Many Y chromosome genes expand in multi-copy families and some evolve under purifying selection. Thus, the Y chromosome exhibits dynamic evolution, whereas the X chromosome is more stable. Mapping short-read sequencing data to these assemblies revealed diversity and selection patterns on sex chromosomes of more than 100 individual great apes. These reference assemblies are expected to inform human evolution and conservation genetics of non-human apes, all of which are endangered species.

Co-opting regulation bypass repair as a gene-correction strategy for monogenic diseases.

With the development of CRISPR-Cas9-mediated gene-editing technologies, correction of disease-causing mutations has become possible. However, current gene-correction strategies preclude mutation repair in post-mitotic cells of human tissues, and a unique repair strategy must be designed and tested for each and every mutation that may occur in a gene. We have developed a novel gene-correction strategy, co-opting regulation bypass repair (CRBR), which can repair a spectrum of mutations in mitotic or post-mitotic cells and tissues. CRBR utilizes the non-homologous end joining (NHEJ) pathway to insert a coding sequence (CDS) and transcription/translation terminators targeted upstream of any CDS mutation and downstream of the transcriptional promoter. CRBR results in simultaneous co-option of the endogenous regulatory region and bypass of the genetic defect. We validated the CRBR strategy for human gene therapy by rescuing a mouse model of Wolcott-Rallison syndrome (WRS) with permanent neonatal diabetes caused by either a large deletion or a nonsense mutation in the PERK (EIF2AK3) gene. Additionally, we integrated a CRBR GFP-terminator cassette downstream of the human insulin promoter in cadaver pancreatic islets of Langerhans, which resulted in insulin promoter regulated expression of GFP, demonstrating the potential utility of CRBR in human tissue gene repair.

The protein kinase PERK/EIF2AK3 regulates proinsulin processing not via protein synthesis but by controlling endoplasmic reticulum chaperones.

Loss-of-function mutations of the protein kinase PERK (EIF2AK3) in humans and mice cause permanent neonatal diabetes and severe proinsulin aggregation in the endoplasmic reticulum (ER), highlighting the essential role of PERK in insulin production in pancreatic ß cells. As PERK is generally known as a translational regulator of the unfolded protein response (UPR), the underlying cause of these ß cell defects has often been attributed to derepression of proinsulin synthesis, resulting in proinsulin overload in the ER. Using high-resolution imaging and standard protein fractionation and immunological methods we have examined the PERK-dependent phenotype more closely. We found that whereas proinsulin aggregation requires new protein synthesis, global protein and proinsulin synthesis are down-regulated in PERK-inhibited cells, strongly arguing against proinsulin overproduction being the root cause of their aberrant ER phenotype. Furthermore, we show that PERK regulates proinsulin proteostasis by modulating ER chaperones, including BiP and ERp72. Transgenic overexpression of BiP and BiP knockdown (KD) both promoted proinsulin aggregation, whereas ERp72 overexpression and knockdown rescued it. These findings underscore the importance of ER chaperones working in concert to achieve control of insulin production and identify a role for PERK in maintaining a functional balance among these chaperones.

Insulin secretion and Ca2+ dynamics in ß-cells are regulated by PERK (EIF2AK3) in concert with calcineurin.

Protein kinase R (PKR)-like endoplasmic reticulum kinase (PERK) (EIF2AK3) is essential for normal development and function of the insulin-secreting ß-cell. Although genetic ablation of PERK in ß-cells results in permanent neonatal diabetes in humans and mice, the underlying mechanisms remain unclear. Here, we used a newly developed and highly specific inhibitor of PERK to determine the immediate effects of acute ablation of PERK activity. We found that inhibition of PERK in human and rodent ß-cells causes a rapid inhibition of secretagogue-stimulated subcellular Ca(2+) signaling and insulin secretion. These dysfunctions stem from alterations in store-operated Ca(2+) entry and sarcoplasmic endoplasmic reticulum Ca(2+)-ATPase activity. We also found that PERK regulates calcineurin, and pharmacological inhibition of calcineurin results in similar defects on stimulus-secretion coupling. Our findings suggest that interplay between calcineurin and PERK regulates ß-cell Ca(2+) signaling and insulin secretion, and that loss of this interaction may have profound implications in insulin secretion defects associated with diabetes.

Do Thai parents discuss sex and AIDS with young adolescents? A qualitative study.

This qualitative study explored parents' and young adolescents' perceptions of communication related to sex and HIV/AIDS. Focus group discussions and group discussion were conducted among 67 adolescents and 30 parents. For the adolescents, group discussion using participatory activities was conducted, followed by five focus group discussions. Group discussions using participatory activities were conducted among parents. Thematic analysis indicated that the adolescents received inadequate information about sex and AIDS from their parents, whom they feared as providing negative judgment, and this represented a key barrier to such discussions. Their parents, on the other hand, reported that they believed their children were still too young to learn about and engage in sexual activities. The parents perceived barriers to communication included a lack of confidence and feelings of embarrassment. Nevertheless, they also recognized their important role in their child's sexual education. Collectively, these results draw attention to the need for a culturally appropriate program to strengthen parent-child communication skills for the topics of sex and HIV/AIDS.

In vivo detection of DNA secondary structures using permanganate/S1 footprinting with direct adapter ligation and sequencing (PDAL-Seq).

DNA secondary structures are essential elements of the genomic landscape, playing a critical role in regulating various cellular processes. These structures refer to G-quadruplexes, cruciforms, Z-DNA or H-DNA structures, amongst others (collectively called 'non-B DNA'), which DNA molecules can adopt beyond the B conformation. DNA secondary structures have significant biological roles, and their landscape is dynamic and can rearrange due to various factors, including changes in cellular conditions, temperature, and DNA-binding proteins. Understanding this dynamic nature is crucial for unraveling their functions in cellular processes. Detecting DNA secondary structures remains a challenge. Conventional methods, such as gel electrophoresis and chemical probing, have limitations in terms of sensitivity and specificity. Emerging techniques, including next-generation sequencing and single-molecule approaches, offer promise but face challenges since these techniques are mostly limited to only one type of secondary structure. Here we describe an updated version of a technique permanganate/S1 nuclease footprinting, which uses potassium permanganate to trap single-stranded DNA regions as found in many non-B structures, in combination with S1 nuclease digest and adapter ligation to detect genome-wide non-B formation. To overcome technical hurdles, we combined this method with direct adapter ligation and sequencing (PDAL-Seq). Furthermore, we established a user-friendly pipeline available on Galaxy to standardize PDAL-Seq data analysis. This optimized method allows the analysis of many types of DNA secondary structures that form in a living cell and will advance our knowledge of their roles in health and disease.

GCN2 regulates the CCAAT enhancer binding protein beta and hepatic gluconeogenesis.

Mice deficient for general control nondepressible-2 (Gcn2) either globally or specifically in the liver display reduced capacity to maintain glucose homeostasis during fasting, suggesting the hypothesis that GCN2 may regulate gluconeogenesis (GNG), which normally plays a key role maintaining peripheral glucose homeostasis. Gcn2-deficient mice exhibit normal insulin sensitivity and plasma insulin but show reduced GNG when administered pyruvate, a gluconeogenic substrate. The basal expression of phosphoenolpyruvate carboxykinase, a rate-limiting enzyme in GNG, is abnormally elevated in Gcn2 knockout (KO) mice in the fed state but fails to be further induced during fasting. The level of tricarboxylic acid cycle intermediates, including malate and oxaloacetate, and the NADH-to-NAD(+) ratio are perturbed in the liver of Gcn2 KO mice either in the fed or fasted state, which may directly impinge upon GNG. Additionally, the expression of the CCAAT enhancer-binding protein-ß (C/EBPß) in the liver fails to be induced in Gcn2 KO mice after 24 h fasting, and the liver-specific Cebpß KO mice show reduced fasting GNG similar to that seen in Gcn2-deficient mice. Our study demonstrates that GCN2 is important in maintaining GNG in the liver, which is likely to be mediated through regulation of C/EBPß.

Perspectives on provider behaviors: a qualitative study of sexual and gender minorities regarding quality of care.

Lesbian, gay, bisexual, transgender, and queer (LGBTQ) identified patients report receiving substandard care from healthcare providers. They face the fear and disturbing reality of discrimination when accessing health care. Without culturally sensitive treatment, nursing and other health professions do not properly care for this population. Following the recent trend towards awareness and need for inclusion of LGBTQ populations in healthcare, this paper provides a summary of the current literature on the treatment and needs of LGBTQ people and describes focus groups conducted to explore perceptions regarding provider behaviors. It concludes with a list of behaviors that enhance or impede quality care that can serve as a guide for healthcare professionals.

Specificity and sensitivity of the social communication questionnaire lifetime screening tool for autism spectrum disorder in a UK CAMHS service.

INTRODUCTION: The Social Communication Questionnaire is used to identify children and young people (CYP) who may require formal ASD assessment. However, there is a paucity of research on its utility in Children and Adolescent Mental Health Services. This evaluation aimed to determine the sensitivity and specificity of the Social Communication Questionnaire (SCQ) in a UK, Midlands CAMHS service. METHOD: Forty young people (mean age 13.75 years) were screened using the caregiver reported SCQ before completing 'gold standard' assessment. RESULTS: The SCQ had a sensitivity of 80% and a specificity of 25.7%. ROC curve analysis indicated low diagnostic accuracy. Differences in predictive accuracy of SCQ and diagnostic standard were statistically significant (p < 0.0001). CONCLUSION: This evaluation builds on previous research suggesting that the SCQ may not be an efficient screening tool in CAMHS settings.

The Complete Sequence and Comparative Analysis of Ape Sex Chromosomes.

Apes possess two sex chromosomes-the male-specific Y and the X shared by males and females. The Y chromosome is crucial for male reproduction, with deletions linked to infertility. The X chromosome carries genes vital for reproduction and cognition. Variation in mating patterns and brain function among great apes suggests corresponding differences in their sex chromosome structure and evolution. However, due to their highly repetitive nature and incomplete reference assemblies, ape sex chromosomes have been challenging to study. Here, using the state-of-the-art experimental and computational methods developed for the telomere-to-telomere (T2T) human genome, we produced gapless, complete assemblies of the X and Y chromosomes for five great apes (chimpanzee, bonobo, gorilla, Bornean and Sumatran orangutans) and a lesser ape, the siamang gibbon. These assemblies completely resolved ampliconic, palindromic, and satellite sequences, including the entire centromeres, allowing us to untangle the intricacies of ape sex chromosome evolution. We found that, compared to the X, ape Y chromosomes vary greatly in size and have low alignability and high levels of structural rearrangements. This divergence on the Y arises from the accumulation of lineage-specific ampliconic regions and palindromes (which are shared more broadly among species on the X) and from the abundance of transposable elements and satellites (which have a lower representation on the X). Our analysis of Y chromosome genes revealed lineage-specific expansions of multi-copy gene families and signatures of purifying selection. In summary, the Y exhibits dynamic evolution, while the X is more stable. Finally, mapping short-read sequencing data from >100 great ape individuals revealed the patterns of diversity and selection on their sex chromosomes, demonstrating the utility of these reference assemblies for studies of great ape evolution. These complete sex chromosome assemblies are expected to further inform conservation genetics of nonhuman apes, all of which are endangered species.

PERK EIF2AK3 control of pancreatic beta cell differentiation and proliferation is required for postnatal glucose homeostasis.

Mutations in PERK (EIF2AK3) result in permanent neonatal diabetes as well as several other anomalies that underlie the human Wolcott-Rallison syndrome, and these anomalies are mirrored in Perk knockout mice. To identify the cause of diabetes in PERK-deficient mice, we generated a series of tissue- and cell-specific knockouts of the Perk gene and performed a developmental analysis of the progression to overt diabetes. We discovered that PERK is specifically required in the insulin-secreting beta cells during the fetal and early neonatal period as a prerequisite for postnatal glucose homeostasis. However, PERK expression in beta cells is not required at the adult stage to maintain beta cell functions and glucose homeostasis. We show that PERK-deficient mice exhibit severe defects in fetal/neonatal beta cell proliferation and differentiation, resulting in low beta cell mass, defects in proinsulin trafficking, and abrogation of insulin secretion that culminate in permanent neonatal diabetes.

Health care transitions among youth with disabilities or special health care needs: an ecological approach.

This literature review of 46 articles uses the ecological model as a framework for organizing concepts and themes related to health care transition among youth with disabilities or special health care needs (SHCN). Transition involves interactions in immediate and distal environmental systems. Important interactions in immediate environments include those with family members, health care providers, and peers. Activities in distal systems include policies at the governmental and health system levels. The ecological model can help researchers and practitioners to design experimental interventions in multiple settings that ensure smooth transitions and support the well-being of youth with disabilities or SHCN.

Creating Project Talanoa: a culturally based community health program for U.S. Pacific Islander adolescents.

OBJECTIVE: This is an evaluation of the process and outcome of a research study to determine a culturally targeted health promotion program for U.S. Pacific Islander youth who are at risk for co-occurring problem behaviors, including risky sexual behavior, substance abuse, and interpersonal violence. DESIGN AND SAMPLE: An exploratory design was used and included qualitative interviews (N=54), focus groups (N=16), participant observation (over 3 years), and surveys (N=24) with Pacific Islander adults and youth. After identifying key cultural values and reviewing existing evidence-based prevention interventions, "Project Talanoa" was developed around 4 constructs: (1) cultural identity and pride, (2) teen health, (3) peer relations, and (4) family ties. The program was pilot tested and evaluated by 24 Pacific Islander adolescents (ages 12-15 years). RESULTS: Results indicate it was culturally appropriate, well liked by the participants, supported by parents and others in the community, and found to be feasible. CONCLUSIONS: Additional research is needed to test it for effectiveness. Project Talanoa provides a model for applying cultural concepts in the development of a risk reduction intervention for adolescents.

Genetic connectivity and population structure of African savanna elephants (Loxodonta africana) in Tanzania.

Increasing human population growth, exurban development, and associated habitat fragmentation is accelerating the isolation of many natural areas and wildlife populations across the planet. In Tanzania, rapid and ongoing habitat conversion to agriculture has severed many of the country's former wildlife corridors between protected areas. To identify historically linked protected areas, we investigated the genetic structure and gene flow of African savanna elephants in Tanzania using microsatellite and mitochondrial DNA markers in 688 individuals sampled in 2015 and 2017. Our results indicate distinct population genetic structure within and between ecosystems across Tanzania, and reveal important priority areas for connectivity conservation. In northern Tanzania, elephants sampled from the Tarangire-Manyara ecosystem appear marginally, yet significantly isolated from elephants sampled from the greater Serengeti ecosystem (mean F ST = 0.03), where two distinct subpopulations were identified.Unexpectedly, elephants in the Lake Manyara region appear to be more closely related to those across the East African Rift wall in the Ngorongoro Conservation Area than they are to the neighboring Tarangire subpopulations. We concluded that the Rift wall has had a negligible influence on genetic differentiation up to this point, but differentiation may accelerate in the future because of ongoing loss of corridors in the area. Interestingly, relatively high genetic similarity was found between elephants in Tarangire and Ruaha although they are separated by >400 km. In southern Tanzania, there was little evidence of female-mediated gene flow between Ruaha and Selous, probably due to the presence of the Udzungwa Mountains between them. Despite observing evidence of significant isolation, the populations of elephants we examined generally exhibited robust levels of allelic richness (mean A R = 9.96), heterozygosity (mean µH E = 0.73), and effective population sizes (mean N e = 148). Our results may inform efforts to restore wildlife corridors between protected areas in Tanzania in order to facilitate gene flow for long-term survival of elephants and other species.

Acute ablation of PERK results in ER dysfunctions followed by reduced insulin secretion and cell proliferation.

BACKGROUND: A deficiency in Perk (EIF2AK3) causes multiple neonatal defects in humans known as the Wolcott Rallison syndrome. Perk KO mice exhibit the same array of defects including permanent neonatal diabetes (PND). PND in mice was previously shown by us to be due to a decrease in beta cell proliferation and insulin secretion. The aim of this study was to determine if acute ablation of PERK in the 832/13 beta cells recapitulates these defects and to identify the primary molecular basis for beta cell dysfunction. RESULTS: The INS1 832/13 transformed rat beta cell line was transduced with a dominant-negative Perk transgene via an adenoviral vector. AdDNPerk-832/13 beta cells exhibited reduced expression of insulin and MafA mRNAs, reduced insulin secretion, and reduced cell proliferation. Although proinsulin content was reduced in AdDNPerk-832/13 beta cells, proinsulin was abnormally retained in the endoplasmic reticulum. A temporal study of the acute ablation of Perk revealed that the earliest defect seen was induced expression of two ER chaperone proteins, GRP78/BiP and ERp72. The oxidized states of ERp72 and ERp57 were also increased suggesting an imbalance in the redox state of the ER. CONCLUSION: Acute ablation of Perk in INS 832/13 beta cells exhibited all of the major defects seen in Perk KO mice and revealed abnormal expression and redox state of key ER chaperone proteins. Dysregulation of ER chaperone/folding enzymes ERp72 and GRP78/BiP occurred early after ablation of PERK function suggesting that changes in ER secretory functions may give rise to the other defects including reduced insulin gene expression, secretion, and cell proliferation.

When family means more (or less) than genetics: the intersection of culture, family and genomics.

There is renewed interest in the use of family history to predict individual disease susceptibility, and as a result, standardized online family history tools are being developed and marketed as a "new genetic test." It is not known how cultural variations in definitions of family influence collection of these data or what is the best format to use. This is significant given that the populations who carry the greatest burden of the target diseases have not been considered in efforts to test these tools. A qualitative study with a convenience sample of 19 Japanese Americans and Samoan Americans, two groups at high risk for type 2 diabetes, was conducted to explore the process of collecting family history. A particularly strong finding was the high degree of acceptance experienced by the participants with the process and their pride in visualizing their family graphically displayed in pedigrees. It was also found that Samoans included those linked by nonbiological ties in their families, which reflects their cultural practices. Further research is needed to assess the most effective and efficient way to gather family history given the complexities surrounding the deceptively simple concept of family.

From manageable to losing control: a grounded theory study of psychosis risk syndrome.

AIM: The purpose of this study is to develop a theoretical explanation of the prodromal schizophrenia process, or so-called psychosis risk syndrome, by describing patients' own experiences with symptoms, thoughts and feelings. METHODS: A total of 40 interviews were conducted in Taiwan. A Grounded Theory method was selected because of its demonstrated effectiveness in generating theory around dynamic and complex processes on which little is known, all of which is the case with psychosis risk syndrome. Constant comparison analysis, memo writing, member checking, and theoretical sampling were adopted. RESULTS: A core theoretical framework was developed in which the process of the psychosis risk syndrome is described as proceeding from manageable to uncontrollable. Four stages emerged from the analysis: (1) something is wrong, (2) boiling up, (3) breaking point, and (4) losing control. CONCLUSIONS: The framework resulting from this Grounded Theory research is innovative in presenting patterns and clinical staging that marks the progression from premorbid stage to full-blown psychosis. In addition to specifying the detailed process through in-depth interviews, this research makes two fundamental contributions by: (1) adding evidence to current science and (2) taking patients' experience into consideration to improve the validity of screening tools and design appropriate intervention programs for people with early warning signs of developing schizophrenia.

PERK is essential for neonatal skeletal development to regulate osteoblast proliferation and differentiation.

Loss of function mutations of Perk (eukaryotic translation initiation factor 2 alpha kinase 3) in humans and mice cause severe neonatal developmental defects, including diabetes, growth retardation and multiple skeletal dysplasias. Comprehensive analyses on bone tissue, at the cellular and molecular level in PERK-deficient mice demonstrated that neonatal Perk-/- mice are severely osteopenic, which is caused by a deficiency in the number of mature osteoblasts, impaired osteoblast differentiation, and reduced type I collagen secretion. Impaired differentiation of osteoblasts in Perk KO mice was associated with decreased expression of Runx2 and Osterix, key regulators of osteoblast development. Reduced cell proliferation and reduced expression of key cell cycle factors including cyclin D, cyclin E, cyclin A, Cdc2, and CDK2 occur in parallel with the differentiation defect in mutant osteoblasts. In addition, the trafficking and secretion of type I collagen is compromised as manifested by abnormal retention of procollagen I in the endoplasmic reticulum, and reduced mature collagen production and mineralization. Taken together, these studies identify PERK as a novel regulator of skeletal development and osteoblast biology.

An approach to understanding the interaction of hope and desire for explicit prognostic information among individuals with severe chronic obstructive pulmonary disease or advanced cancer.

BACKGROUND: Physicians often report that they are reluctant to discuss prognosis for life-threatening illnesses with patients and family out of concern for destroying their hope, yet there is little empirical research describing how patients and family incorporate their needs for hope with desires for prognostic information. OBJECTIVE: We conducted a qualitative study to examine the perspectives of patients, family, physicians, and nurses on the simultaneous need for supporting hope and discussing prognosis. METHODS: We conducted in-depth longitudinal qualitative interviews with patients with either advanced cancer or severe chronic obstructive pulmonary disease (COPD), along with their family, physicians, and nurses. We used principles of grounded theory to analyze the transcripts and evaluated a conceptual model with four diagrams depicting different types of approaches to hope and prognostic information. RESULTS: We interviewed 55 patients, 36 family members, 31 physicians, and 25 nurses representing 220 hours of interviews. Asking patients directly "how much information" they wanted was, by itself, not useful for identifying information needs, but in-depth questioning identified variability in patients' and family members' desires for explicit prognostic information. All but 2 patients endorsed at least one of the diagrams concerning the interaction of hope and prognostic information and some patients described moving from one diagram to another over the course of their illness. Respondents also described two different approaches to communication about prognosis based on the diagram selected: two of the four diagrams suggested a direct approach and the other two suggested a cautious, indirect approach. CONCLUSIONS: This study found important variability in the ways different patients with life-limiting illnesses approach the interaction of wanting support for hope and prognostic information from their clinicians. The four-diagram approach may help clinicians understand individual patients and families, but further research is needed to determine the utility of these diagrams for improving communication about end-of-life care.

The role of centers in fostering interdisciplinary research.

This article describes the history of the University of Washington School of Nursing (UW-SON) Center for Women's Health and Gender Research (CWHGR) and its role in helping initiate and sustain interdisciplinary research. The growing focus on the need for interdisciplinary research has made it imperative that nursing scientists collaborate with colleagues in other health-related fields including medicine, public health, dentistry, and social work. The CWHGR increased interdisciplinary research activities through mentorship of faculty and trainees, the creation of core laboratory facilities, the award of pilot grant funding focused on interdisciplinary collaboration, consultation on research design and methods both within and outside the UW-SON, and the utilization of the Human Response Model for both biobehavioral and sociocultural research collaboration. Accomplishments as well as lessons learned related to interdisciplinary research during the 19 years of the UW-SON CWHGR are highlighted.