Structural alterations in the amygdala and impaired social incentive learning in a mouse model of a genetic variant associated with neurodevelopmental disorders.

Copy number variants (CNVs) are robustly associated with psychiatric disorders and their dimensions and changes in brain structures and behavior. However, as CNVs contain many genes, the precise gene-phenotype relationship remains unclear. Although various volumetric alterations in the brains of 22q11.2 CNV carriers have been identified in humans and mouse models, it is unknown how the genes in the 22q11.2 region individually contribute to structural alterations and associated mental illnesses and their dimensions. Our previous studies have identified Tbx1, a T-box family transcription factor encoded in 22q11.2 CNV, as a driver gene for social interaction and communication, spatial and working memory, and cognitive flexibility. However, it remains unclear how TBX1 impacts the volumes of various brain regions and their functionally linked behavioral dimensions. In this study, we used volumetric magnetic resonance imaging analysis to comprehensively evaluate brain region volumes in congenic Tbx1 heterozygous mice. Our data show that the volumes of anterior and posterior portions of the amygdaloid complex and its surrounding cortical regions were reduced in Tbx1 heterozygous mice. Moreover, we examined the behavioral consequences of an altered volume of the amygdala. Tbx1 heterozygous mice were impaired for their ability to detect the incentive value of a social partner in a task that depends on the amygdala. Our findings identify the structural basis for a specific social dimension associated with loss-of-function variants of TBX1 and 22q11.2 CNV.

Cognition and dementia with Raymond and Brain: Curriculum development and evaluation using interactive animated flipped-classroom modules to impact nursing students' attitude toward dementia care.

AIM: To design a modular, flipped-classroom curriculum using character animations to improve knowledge and attitudes regarding dementia care among pre-clinical nursing students. BACKGROUND: Demographic trends suggest an urgent, unmet need for nurses with interest and adequate training in caring for people with dementia and other disorders of cognition. While flipped classrooms using video show promise, little is known about specific animation techniques to impact knowledge and attitudes in preclinical nursing education. DESIGN: A curriculum was developed, implemented and assessed across three nursing schools in series, totaling 223 eligible students in the states of Connecticut and Hawaii, USA from 2019 to 2022. The evaluation included prospective pre-post assessment of knowledge, attitudes and module acceptability, as well as qualitative interpretations of needs assessment data. METHODS: The six-step curriculum development process was based on that described by Kern et al., including: 1) general needs assessment in the form of literature review; 2) targeted needs assessment, in the form of faculty stakeholder meetings, a student focus group and baseline surveys; 3) optimization of learning objectives based on needs; 4) development of a pedagogical approach, namely animated, interactive modules informed by previously described best practices in animation development; 5) implementation across three different nursing schools; and 6) assessment of the learners and evaluation of the curriculum, primarily via surveys and engagement metadata. RESULTS: Needs assessments confirmed the importance of prior experiences, sense of mission and other affective elements as key factors mitigating learners' baseline receptiveness to training and careers in cognition-related care. Students at all three institutions rated the modules' impact on their dementia-related attitudes highly, however these ratings were statistically significantly lower when both modules were delivered as a single assignment at one site. Knowledge quiz scores significantly increased from baseline at all three sites. Only 2.6% of respondents would have preferred a text-based reading assignment. Acceptability scores, including clarity, relevance, entertainment, attention and complexity, were generally rated highly, but attention and entertainment were rated significantly lower when both modules were administered as a single assignment. CONCLUSION: Cognition and Dementia with Raymond and Brain demonstrates the successful blending of animation industry workflows with best practices of curriculum development to create a novel, animated module series that is acceptable and effective in priming nursing students with the attitudes and knowledge to continue learning about cognition and its disorders.

Structural alterations in the amygdala and impaired social incentive learning in a mouse model of a genetic variant associated with neurodevelopmental disorders.

Copy number variants (CNVs) are robustly associated with psychiatric disorders and their dimensions and changes in brain structures and behavior. However, as CNVs contain many genes, the precise gene-phenotype relationship remains unclear. Although various volumetric alterations in the brains of 22q11.2 CNV carriers have been identified in humans and mouse models, it is unknown how the genes in the 22q11.2 region individually contribute to structural alterations and associated mental illnesses and their dimensions. Our previous studies have identified Tbx1, a T-box family transcription factor encoded in 22q11.2 CNV, as a driver gene for social interaction and communication, spatial and working memory, and cognitive flexibility. However, it remains unclear how TBX1 impacts the volumes of various brain regions and their functionally linked behavioral dimensions. In this study, we used volumetric magnetic resonance imaging analysis to comprehensively evaluate brain region volumes in congenic Tbx1 heterozygous mice. Our data show that the volumes of anterior and posterior portions of the amygdaloid complex and its surrounding cortical regions were reduced in Tbx1 heterozygous mice. Moreover, we examined the behavioral consequences of an altered volume of the amygdala. Tbx1 heterozygous mice were impaired for their ability to detect the incentive value of a social partner in a task that depends on the amygdala. Our findings identify the structural basis for a specific social dimension associated with loss-of-function variants of TBX1 and 22q11.2 CNV.

Tbx1, a gene encoded in 22q11.2 copy number variant, is a link between alterations in fimbria myelination and cognitive speed in mice.

Copy number variants (CNVs) have provided a reliable entry point to identify the structural correlates of atypical cognitive development. Hemizygous deletion of human chromosome 22q11.2 is associated with impaired cognitive function; however, the mechanisms by which the CNVs contribute to cognitive deficits via diverse structural alterations in the brain remain unclear. This study aimed to determine the cellular basis of the link between alterations in brain structure and cognitive functions in mice with a heterozygous deletion of Tbx1, one of the 22q11.2-encoded genes. Ex vivo whole-brain diffusion-tensor imaging (DTI)-magnetic resonance imaging (MRI) in Tbx1 heterozygous mice indicated that the fimbria was the only region with significant myelin alteration. Electron microscopic and histological analyses showed that Tbx1 heterozygous mice exhibited an apparent absence of large myelinated axons and thicker myelin in medium axons in the fimbria, resulting in an overall decrease in myelin. The fimbria of Tbx1 heterozygous mice showed reduced mRNA levels of Ng2, a gene required to produce oligodendrocyte precursor cells. Moreover, postnatal progenitor cells derived from the subventricular zone, a source of oligodendrocytes in the fimbria, produced fewer oligodendrocytes in vitro. Behavioral analyses of these mice showed selectively slower acquisition of spatial memory and cognitive flexibility with no effects on their accuracy or sensory or motor capacities. Our findings provide a genetic and cellular basis for the compromised cognitive speed in patients with 22q11.2 hemizygous deletion.

Older immigrants perceived health after migration to the United States: Influence of age and level of acculturation.

OBJECTIVES: To determine the age-related differences in the association between level of acculturation and perceptions about change in health status after migration in mid- to late-life immigrants. DESIGN: Cross-sectional study. SETTING: Nationally representative cohort of legal U.S. immigrants. PARTICIPANTS: Legal immigrants ≥50 years of age. MEASUREMENTS: The outcome was perceived change in health status, determined by the survey question, "compared with your health right before you most recently came to the United States to live, would you say that your health is better now, about the same or worse?" The main predictor included age group (50-64, 65-74, and ≥75 years) and secondary variable of interest was level of acculturation. Analyses were adjusted for demographic characteristics, medical, and functional comorbidities. RESULTS: Immigrants age ≥75 years were more likely to report worse health after migration [RRR 1.93, 95% CI (1.17, 3.17), p < 0.01] compared with immigrants of 50-64 years, but this difference was not statistically significant in the adjusted model. Acculturation level was associated with increased likelihood of reporting worse health status, [RRR 2.10, (1.02, 4.35), p < 0.05] for somewhat acculturated and [RRR 2.55, (1.10, 5.88), p < 0.05] for most acculturated, compared with participants who were not acculturated. CONCLUSION: The oldest immigrant group (≥75 years) was more likely to report worse health after migration, but this association was no longer significant after accounting for acculturation level and other covariates. Future work should be undertaken to identify specific health needs across older immigrant age groups and identify acculturative stressors that negatively impact health.

Computational identification of variables in neonatal vocalizations predictive for postpubertal social behaviors in a mouse model of 16p11.2 deletion.

Autism spectrum disorder (ASD) is often signaled by atypical cries during infancy. Copy number variants (CNVs) provide genetically identifiable cases of ASD, but how early atypical cries predict a later onset of ASD among CNV carriers is not understood in humans. Genetic mouse models of CNVs have provided a reliable tool to experimentally isolate the impact of CNVs and identify early predictors for later abnormalities in behaviors relevant to ASD. However, many technical issues have confounded the phenotypic characterization of such mouse models, including systematically biased genetic backgrounds and weak or absent behavioral phenotypes. To address these issues, we developed a coisogenic mouse model of human proximal 16p11.2 hemizygous deletion and applied computational approaches to identify hidden variables within neonatal vocalizations that have predictive power for postpubertal dimensions relevant to ASD. After variables of neonatal vocalizations were selected by least absolute shrinkage and selection operator (Lasso), random forest, and Markov model, regression models were constructed to predict postpubertal dimensions relevant to ASD. While the average scores of many standard behavioral assays designed to model dimensions did not differentiate a model of 16p11.2 hemizygous deletion and wild-type littermates, specific call types and call sequences of neonatal vocalizations predicted individual variability of postpubertal reciprocal social interaction and olfactory responses to a social cue in a genotype-specific manner. Deep-phenotyping and computational analyses identified hidden variables within neonatal social communication that are predictive of postpubertal behaviors.

Maternal approach behaviors toward neonatal calls are impaired by mother's experiences of raising pups with a risk gene variant for autism.

How the intrinsic sequence structure of neonatal mouse pup ultrasonic vocalization (USV) and maternal experiences determine maternal behaviors in mice is poorly understood. Our previous work showed that pups with a Tbx1 heterozygous (HT) mutation, a genetic risk for autism spectrum disorder (ASD), emit altered call sequences that do not induce maternal approach behaviors in C57BL6/J mothers. Here, we tested how maternal approach behaviors induced by wild-type and HT USVs are influenced by the mother's experience in raising pups of these two genotypes. The results showed that wild-type USVs were effective in inducing maternal approach behaviors when mothers raised wild-type but not HT pups. The USVs of HT pups were ineffective regardless of whether mothers raised HT or wild-type pups. However, the sequence structure of pup USVs had no effect on the general, non-directional incentive motivation of maternal behaviors. Our data show how the mother's experience with a pup with a genetic risk for ASD alters the intrinsic incentive values of USV sequences in maternal approach behaviors.

Heterozygosity of murine Crkl does not recapitulate behavioral dimensions of human 22q11.2 hemizygosity.

Deletions in 22q11.2 human chromosome are known to be associated with psychiatric disorders, such as intellectual disability, schizophrenia, autism spectrum disorder, and anxiety disorders. This copy number variation includes a 3.0 Mb deletion and a nested proximal 1.5 Mb hemizygous deletion in the same region. Evidence indicates that the distal 22q11.2 region outside the nested 1.5 Mb deletion also might be contributory in humans. However, the precise genetic architecture within the distal region responsible for psychiatric disorders remains unclear, and this issue cannot be experimentally evaluated beyond the correlation in humans. As CRKL (CRK-like Proto-Oncogene, Adaptor Protein) is one of the genes encoded in the distal 22q11.2 segment and its homozygous deletion causes physical phenotypes of 22q11.2 hemizygous deletion, we tested the hypothesis that its murine homolog Crkl contributes to behavioral phenotypes relevant to psychiatric disorders in mice. Congenic Crkl heterozygosity reduced thigmotaxis, an anxiety-related behavior, in an inescapable open field, but had no apparent effect on social interaction, spontaneous alternation in a T-maze, anxiety-like behavior in an elevated plus maze, or motor activity in an open field. Our data indicate that the heterozygosity of murine Crkl does not recapitulate social deficits, working memory deficits, repetitive behavior traits or hyperactivity of human 22q11.2 hemizygous deletion. Moreover, while 22q11.2 hemizygous deletion is associated with high levels of phobia and anxiety in humans, our data suggest that Crkl heterozygosity rather acts as a protective factor for phobia-like behavior in an open field.

Presynaptic Vesicle Protein SEPTIN5 Regulates the Degradation of APP C-Terminal Fragments and the Levels of Aß.

Alzheimer's disease (AD) is a neurodegenerative disease characterized by aberrant amyloid-ß (Aß) and hyperphosphorylated tau aggregation. We have previously investigated the involvement of SEPTIN family members in AD-related cellular processes and discovered a role for SEPTIN8 in the sorting and accumulation of ß-secretase. Here, we elucidated the potential role of SEPTIN5, an interaction partner of SEPTIN8, in the cellular processes relevant for AD, including amyloid precursor protein (APP) processing and the generation of Aß. The in vitro and in vivo studies both revealed that the downregulation of SEPTIN5 reduced the levels of APP C-terminal fragments (APP CTFs) and Aß in neuronal cells and in the cortex of Septin5 knockout mice. Mechanistic elucidation revealed that the downregulation of SEPTIN5 increased the degradation of APP CTFs, without affecting the secretory pathway-related trafficking or the endocytosis of APP. Furthermore, we found that the APP CTFs were degraded, to a large extent, via the autophagosomal pathway and that the downregulation of SEPTIN5 enhanced autophagosomal activity in neuronal cells as indicated by altered levels of key autophagosomal markers. Collectively, our data suggest that the downregulation of SEPTIN5 increases the autophagy-mediated degradation of APP CTFs, leading to reduced levels of Aß in neuronal cells.

Association of Patient Priorities-Aligned Decision-Making With Patient Outcomes and Ambulatory Health Care Burden Among Older Adults With Multiple Chronic Conditions: A Nonrandomized Clinical Trial.

Importance: Health care may be burdensome and of uncertain benefit for older adults with multiple chronic conditions (MCCs). Aligning health care with an individual's health priorities may improve outcomes and reduce burden. Objective: To evaluate whether patient priorities care (PPC) is associated with a perception of more goal-directed and less burdensome care compared with usual care (UC). Design, Setting, and Participants: Nonrandomized clinical trial with propensity adjustment conducted at 1 PPC and 1 UC site of a Connecticut multisite primary care practice that provides care to almost 15% of the state's residents. Participants included 163 adults aged 65 years or older who had 3 or more chronic conditions cared for by 10 primary care practitioners (PCPs) trained in PPC and 203 similar patients who received UC from 7 PCPs not trained in PPC. Participant enrollment occurred between February 1, 2017, and March 31, 2018; follow-up extended for up to 9 months (ended September 30, 2018). Interventions: Patient priorities care, an approach to decision-making that includes patients' identifying their health priorities (ie, specific health outcome goals and health care preferences) and clinicians aligning their decision-making to achieve these health priorities. Main Outcomes and Measures: Primary outcomes included change in patients' Older Patient Assessment of Chronic Illness Care (O-PACIC), CollaboRATE, and Treatment Burden Questionnaire (TBQ) scores; electronic health record documentation of decision-making based on patients' health priorities; medications and self-management tasks added or stopped; and diagnostic tests, referrals, and procedures ordered or avoided. Results: Of the 366 patients, 235 (64.2%) were female and 350 (95.6%) were white. Compared with the UC group, the PPC group was older (mean [SD] age, 74.7 [6.6] vs 77.6 [7.6] years) and had lower physical and mental health scores. At follow-up, PPC participants reported a 5-point greater decrease in TBQ score than those who received UC (ß [SE], -5.0 [2.04]; P = .01) using a weighted regression model with inverse probability of PCP assignment weights; no differences were seen in O-PACIC or CollaboRATE scores. Health priorities-based decisions were mentioned in clinical visit notes for 108 of 163 (66.3%) PPC vs 0 of 203 (0%) UC participants. Compared with UC patients, PPC patients were more likely to have medications stopped (weighted comparison, 52.0% vs 33.8%; adjusted odds ratio [AOR], 2.05; 95% CI, 1.43-2.95) and less likely to have self-management tasks (57.5% vs 62.1%; AOR, 0.59; 95% CI, 0.41-0.84) and diagnostic tests (80.8% vs 86.4%; AOR, 0.22; 95% CI, 0.12-0.40) ordered. Conclusions and Relevance: This study's findings suggest that patient priorities care may be associated with reduced treatment burden and unwanted health care. Care aligned with patients' priorities may be feasible and effective for older adults with MCCs. Trial Registration: ClinicalTrials.gov identifier: NCT03600389.

Computational Analysis of Neonatal Mouse Ultrasonic Vocalization.

Neonatal vocalization is structurally altered in mouse models of autism spectrum disorder (ASD). Our published data showed that pup vocalization, under conditions of maternal separation, contains sequences whose alterations in a genetic mouse model of ASD impair social communication between pups and mothers. We describe details of a method which reveals the statistical structure of call sequences that are functionally critical for optimal maternal care. Entropy analysis determines the degree of non-random call sequencing. A Markov model determines the actual call sequences used by pups. Sparse partial least squares discriminant analysis (sPLS-DA) identifies call sequences that differentiate groups and reveals the degrees of individual variability in call sequences between groups. These three sets of analyses can be used to identify the otherwise hidden call structure that is altered in mouse models of developmental neuropsychiatric disorders, including not only autism but also schizophrenia. © 2018 by John Wiley & Sons, Inc.

Pre-admission functional status impacts the performance of the APACHE IV model of mortality prediction in critically ill patients.

BACKGROUND: Functional status (FS) before intensive care unit (ICU) admission is associated with short-term and long-term outcomes among critically ill patients. However, measures of FS are generally not integrated into ICU-specific mortality prediction models. METHODS: This retrospective cohort study used prospectively collected data from 9638 consecutive patients admitted to a single ICU between 1 October 2005 and 30 September 2015. For each ICU admission, FS was prospectively determined and classified into three discrete categories based on performance of basic daily living activities (FS1 - fully independent; FS2 - partly dependent; FS3 - completely dependent). We prospectively calculated Acute Physiology and Chronic Health Evaluation (APACHE) IV predicted mortality percentage (APIV PM) for each admission and calculated observed-expected mortality ratios (OEMR), stratified by FS category and APIV PM. We calculated area under the receiver operator characteristic curve (AUC) for APIV PM and mortality for the entire cohort and the three FS categories. RESULTS: Patients had a median (IQR) age of 67 (52-80) years and mean (SD) APIV PM was 18.3% (24.3%). Of these, 7714 (80.0%) were classified as FS1, 1728 (17.9%) as FS2 and 196 (2.0%) as FS3. FS1 patients were younger, had less comorbid disease, and lower APIV PM compared to FS2 and FS3. The OEMR were significantly lower for FS1 (0.67) than FS2 (0.93) or FS3 (0.90) (p < 0.0001 for both comparisons). Among patients with APIV PM 0-10%, 10-25%, 25-50% and ≥50% the OEMR for FS1 were 0.33, 0.49, 0.61 and 0.86. The AUC (95% CI) for APIV PM and mortality for FS1, FS2 and FS3 were 0.924 (0.914-0.933), 0.837 (0.816-0.858) and 0.775 (0.705-0.8456), respectively (p < 0.001 for each comparison). Multivariable analysis demonstrated that FS2 (OR 2.18 (1.84-2.57) (p < 0.0001)) and FS3 (OR 1.99 (1.34-2.96) (p = 0.0006)) were independently associated with increased risk of mortality. CONCLUSIONS: Baseline FS prior to critical illness is a strong independent predictor of mortality and impacts the relationship between observed and APIV PM in those with lower illness severity. Future iterations of mortality prediction models should integrate a baseline measure of FS to improve performance.

Human COMT over-expression confers a heightened susceptibility to dyskinesia in mice.

Catechol-O-methyltransferase (COMT) degrades dopamine and its precursor l-DOPA and plays a critical role in regulating synaptic dopamine actions. We investigated the effects of heightened levels of COMT on dopamine-regulated motor behaviors and molecular alterations in a mouse model of dyskinesia. Transgenic mice overexpressing human COMT (TG) and their wildtype (WT) littermates received unilateral 6-OHDA lesions in the dorsal striatum and were treated chronically with l-DOPA for two weeks. l-DOPA-induced dyskinesia was exacerbated in TG mice without altering l-DOPA motor efficacy as determined by contralateral rotations or motor coordination. Inductions of FosB and phospho-acetylated histone 3 (molecular correlates of dyskinesia) were potentiated in the lesioned striatum of TG mice compared with their WT littermates. The TG mice had lower basal levels of dopamine in the striatum. In mice with lesions, l-DOPA induces a greater increase in the dopamine metabolite 3-methoxytyramine in the lesioned striatum of dyskinetic TG mice than in WT mice. The levels of serotonin and its metabolite were similar in TG and WT mice. Our results demonstrate that human COMT overexpression confers a heightened susceptibility to l-DOPA-induced dyskinesia and alters molecular and neurochemical responses in the lesioned striatum of mice.

Molecular Histochemistry Identifies Peptidomic Organization and Reorganization Along Striatal Projection Units.

Matrix-assisted laser desorption ionization (MALDI) imaging mass spectrometry (IMS) (MALDI-IMS) provides a technical means for simultaneous analysis of precise anatomic localization and regulation of peptides. We explored the technical capability of matrix-assisted laser desorption ionization mass spectrometry for characterization of peptidomic regulation by an addictive substance along two distinct projection systems in the mouse striatum. The spatial expression patterns of substance P and proenkephalin, marker neuropeptides of two distinct striatal projection neurons, were negatively correlated at baseline. We detected 768 mass/charge (m/z) peaks whose expression levels were mostly negatively and positively correlated with expression levels of substance P and proenkephalin A (amino acids 218-228), respectively, within the dorsal striatum. After nicotine administration, there was a positive shift in correlation of mass/charge peak expression levels with substance P and proenkephalin A (218-228). Our exploratory analyses demonstrate the technical capacity of MALDI-IMS for comprehensive identification of peptidomic regulation patterns along histochemically distinguishable striatal projection pathways.

Alterations of social interaction through genetic and environmental manipulation of the 22q11.2 gene Sept5 in the mouse brain.

Social behavior dysfunction is a symptomatic element of schizophrenia and autism spectrum disorder (ASD). Although altered activities in numerous brain regions are associated with defective social cognition and perception, the causative relationship between these altered activities and social cognition and perception-and their genetic underpinnings-are not known in humans. To address these issues, we took advantage of the link between hemizygous deletion of human chromosome 22q11.2 and high rates of social behavior dysfunction, schizophrenia and ASD. We genetically manipulated Sept5, a 22q11.2 gene, and evaluated its role in social interaction in mice. Sept5 deficiency, against a high degree of homogeneity in a congenic genetic background, selectively impaired active affiliative social interaction in mice. Conversely, virally guided overexpression of Sept5 in the hippocampus or, to a lesser extent, the amygdala elevated levels of active affiliative social interaction in C57BL/6J mice. Congenic knockout mice and mice overexpressing Sept5 in the hippocampus or amygdala were indistinguishable from control mice in novelty and olfactory responses, anxiety or motor activity. Moreover, post-weaning individual housing, an environmental condition designed to reduce stress in male mice, selectively raised levels of Sept5 protein in the amygdala and increased active affiliative social interaction in C57BL/6J mice. These findings identify this 22q11.2 gene in the hippocampus and amygdala as a determinant of social interaction and suggest that defective social interaction seen in 22q11.2-associated schizophrenia and ASD can be genetically and environmentally modified by altering this 22q11.2 gene.

Localization of septin proteins in the mouse cochlea.

Septins are a family of GTP binding proteins that are well conserved in eukaryotic species except plants. Septins contribute to the lateral compartmentalization of membranes, cortical rigidity, and the regulation of membrane trafficking by associating with membrane lipids, actin, and microtubules. The organ of Corti in the cochlea has pivotal roles in auditory perception and includes two kinds of highly polarized cells, hair and supporting cells, both of which are rich in actin and microtubules. To identify the roles of septins in the cochlea, we analyzed the localization of three septin proteins, septin 4 (SEPT4), septin 5 (SEPT5), and septin 7 (SEPT7) that are abundantly expressed in brain tissues, and also examined auditory functions of Sept4 and Sept5 null mice. SEPT4, SEPT5, and SEPT7 were expressed in inner and outer pillar cells and Deiters' cells but the distribution patterns of each protein in Deiters' cells were different. SEPT4 and SEPT7 were expressed in the phalangeal process where SEPT5 was not detected. In addition to these cells SEPT5 and SEPT7 were co-localized with presynaptic vesicles of efferent nerve terminals. Only SEPT7 was expressed in the cochlea at embryonic stages. Although expression patterns of septin proteins suggested their important roles in the function of the cochlea, both Sept4 and Sept5 null mice had similar auditory functions to their wild type littermates. Immunohistochemical analysis of Sept4 null mice showed that compensatory expression of SEPT5 in the phalangeal process of Deiters' cells may have caused functional compensation of hearing ability in Sept4 null mice.

Novel features of ARS selection in budding yeast Lachancea kluyveri.

BACKGROUND: The characterization of DNA replication origins in yeast has shed much light on the mechanisms of initiation of DNA replication. However, very little is known about the evolution of origins or the evolution of mechanisms through which origins are recognized by the initiation machinery. This lack of understanding is largely due to the vast evolutionary distances between model organisms in which origins have been examined. RESULTS: In this study we have isolated and characterized autonomously replicating sequences (ARSs) in Lachancea kluyveri - a pre-whole genome duplication (WGD) budding yeast. Through a combination of experimental work and rigorous computational analysis, we show that L. kluyveri ARSs require a sequence that is similar but much longer than the ARS Consensus Sequence well defined in Saccharomyces cerevisiae. Moreover, compared with S. cerevisiae and K. lactis, the replication licensing machinery in L. kluyveri seems more tolerant to variations in the ARS sequence composition. It is able to initiate replication from almost all S. cerevisiae ARSs tested and most Kluyveromyces lactis ARSs. In contrast, only about half of the L. kluyveri ARSs function in S. cerevisiae and less than 10% function in K. lactis. CONCLUSIONS: Our findings demonstrate a replication initiation system with novel features and underscore the functional diversity within the budding yeasts. Furthermore, we have developed new approaches for analyzing biologically functional DNA sequences with ill-defined motifs.

Tbx1: identification of a 22q11.2 gene as a risk factor for autism spectrum disorder in a mouse model.

Although twin studies indicate clear genetic bases of autism spectrum disorder (ASD), the precise mechanisms through which genetic variations causally result in ASD are poorly understood. Individuals with 3 Mb and nested 1.5 Mb hemizygosity of the chromosome 22q11.2 represent genetically identifiable cases of ASD. However, because more than 30 genes are deleted even in the minimal deletion cases of 22q11.2 deficiency, the individual 22q11.2 gene(s) responsible for ASD remain elusive. Here, we examined the impact of constitutive heterozygosity of Tbx1, a 22q11.2 gene, on the behavioral phenotypes of ASD and characterized the regional and cellular expression of its mRNA and protein in mice. Congenic Tbx1 heterozygous (HT) mice were impaired in social interaction, ultrasonic vocalization, memory-based behavioral alternation, working memory and thigmotaxis, compared with wild-type (WT) mice. These phenotypes were not due to non-specific alterations in olfactory function, exploratory behavior, motor movement or anxiety-related behavior. Tbx1 mRNA and protein were ubiquitously expressed throughout the brains of C57BL/6J mice, but protein expression was enriched in regions that postnatally retain the capacity of neurogenesis, and in fact, postnatally proliferating cells expressed Tbx1. In postnatally derived hippocampal culture cells of C57BL/6J mice, Tbx1 levels were higher during proliferation than during differentiation, and expressed in neural progenitor cells, immature and matured neurons and glial cells. Taken together, our data suggest that Tbx1 is a gene responsible for the phenotypes of 22q11.2 hemizygosity-associated ASD possibly through its role in diverse cell types, including postnatally and prenatally generated neurons.

Over-expression of a human chromosome 22q11.2 segment including TXNRD2, COMT and ARVCF developmentally affects incentive learning and working memory in mice.

Duplication of human chromosome 22q11.2 is associated with elevated rates of mental retardation, autism and many other behavioral phenotypes. However, because duplications cover 1.5-6 Mb, the precise manner in which segments of 22q11.2 causally affect behavior is not known in humans. We have now determined the developmental impact of over-expression of an approximately 190 kb segment of human 22q11.2, which includes the genes TXNRD2, COMT and ARVCF, on behaviors in bacterial artificial chromosome (BAC) transgenic (TG) mice. BAC TG mice and wild-type (WT) mice were tested for their cognitive capacities, affect- and stress-related behaviors and motor activity at 1 and 2 months of age. An enzymatic assay determined the impact of BAC over-expression on the activity level of COMT. BAC TG mice approached a rewarded goal faster (i.e. incentive learning), but were impaired in delayed rewarded alternation during development. In contrast, BAC TG and WT mice were indistinguishable in rewarded alternation without delays, spontaneous alternation, prepulse inhibition, social interaction, anxiety-, stress- and fear-related behaviors and motor activity. Compared with WT mice, BAC TG mice had an approximately 2-fold higher level of COMT activity in the prefrontal cortex, striatum and hippocampus. These data suggest that over-expression of this 22q11.2 segment enhances incentive learning and impairs the prolonged maintenance of working memory, but has no apparent effect on working memory per se, affect- and stress-related behaviors or motor capacity. High copy numbers of this 22q11.2 segment might contribute to a highly selective set of phenotypes in learning and cognition during development.

Sept5 deficiency exerts pleiotropic influence on affective behaviors and cognitive functions in mice.

Deletion or duplication of the human chromosome 22q11.2 is associated with many behavioral traits and neuropsychiatric disorders, including autism spectrum disorders and schizophrenia. However, why phenotypes vary widely among individuals with identical deletions or duplications of 22q11.2 and which specific 22q11.2 genes contribute to these phenotypes are still poorly understood. Previous studies have identified a approximately 200 kb 22q11.2 region that contributes to behavioral phenotypes in mice. We tested the role of Septin 5 (Sept5), a gene encoded in the approximately 200 kb region, in affective behaviors, cognitive capacities and motor activity. To evaluate the impact of genetic backgrounds on behavioral phenotypes of Sept5 deficiency, we used mice on two genetic backgrounds. Our data show that Sept5 deficiency decreased affiliative active social interaction, but this phenotypic expression was influenced by genetic backgrounds. In contrast, Sept5 deficiency decreased anxiety-related behavior, increased prepulse inhibition and delayed acquisition of rewarded goal approach, independent of genetic background. These data suggest that Sept5 deficiency exerts pleiotropic effects on a select set of affective behaviors and cognitive processes and that genetic backgrounds could provide an epistatic influence on phenotypic expression.