Allosteric Regulation of Switch-II Domain Controls KRAS Oncogenicity.

RAS proteins are GTPases that regulate a wide range of cellular processes. RAS activity is dependent on its nucleotide-binding status, which is modulated by guanine nucleotide exchange factors (GEF) and GTPase-activating proteins (GAP). KRAS can be acetylated at lysine 104 (K104), and an acetylation-mimetic mutation of K104 to glutamine (K104Q) attenuates the in vitro-transforming capacity of oncogenic KRAS by interrupting GEF-induced nucleotide exchange. To assess the effect of this mutation in vivo, we used CRISPR-Cas9 to generate mouse models carrying the K104Q point mutation in wild-type and conditional KrasLSL-G12D alleles. Homozygous animals for K104Q were viable, fertile, and arose at the expected Mendelian frequency, indicating that K104Q is not a complete loss-of-function mutation. Consistent with our previous findings from in vitro studies, however, the oncogenic activity of KRASG12D was significantly attenuated by mutation at K104. Biochemical and structural analysis indicated that the G12D and K104Q mutations cooperate to suppress GEF-mediated nucleotide exchange, explaining the preferential effect of K104Q on oncogenic KRAS. Furthermore, K104 functioned in an allosteric network with M72, R73, and G75 on the α2 helix of the switch-II region. Intriguingly, point mutation of glycine 75 to alanine (G75A) also showed a strong negative regulatory effect on KRASG12D. These data demonstrate that lysine at position 104 is critical for the full oncogenic activity of mutant KRAS and suggest that modulating the sites in its allosteric network may provide a unique therapeutic approach in cancers expressing mutant KRAS. SIGNIFICANCE: An allosteric network formed by interaction between lysine 104 and residues in the switch-II domain is required for KRAS oncogenicity, which could be exploited for developing inhibitors of the activated oncoprotein.

Neural processes related to negative self-concept in adult and adolescent anorexia nervosa.

OBJECTIVE: Negative self-concept is characteristic of anorexia nervosa (AN), but the neural processes mediating this component of AN is unknown. These studies investigated how valence and social perspectives impact neural processing in both adults and adolescents with AN. METHOD: In an fMRI task, participants evaluated positive and negative adjectives from three social perspectives. Two studies were completed, one in 59 women (healthy, with AN, recovered from AN) and one in 42 adolescents (healthy, with AN). Neural regions of interest (ROIs) related to valence were identified and activations compared across groups and social perspectives. RESULTS: Behaviourally, both adult and adolescent cohorts with AN were less positive during self-evaluations. In the adult study, neural differences related to clinical group and condition were observed in ROIs more responsive to positive social stimuli (medial prefrontal cortex, precuneus, left temporoparietal junction) but not in ROIs more responsive to negative social stimuli. No neural differences in relation to clinical group were observed in the adolescents. CONCLUSIONS: Behavioural differences related to negative self-concept are present in both adolescents and adults with AN, and neural differences, selective for positive social stimuli, were also observed in adults. AN may interfere with neurodevelopmental processes involved in positive self-concept.

Qualitative patient experiences from the Self-Blame and Perspective-Taking Intervention for eating disorders.

BACKGROUND: Problems in social cognition and social support contribute to eating disorders (ED). Group therapy provides an ideal format to create an experiential learning environment focused on understanding social interactions. This pilot study examined the qualitative content of the participants' experiences in the Self-Blame and Perspective-Taking Intervention (SBPI) for ED. METHODS: The SBPI was a 4-week group therapy intervention involving art therapy and psychoeducation that focused on social behaviors in ED patients. Participants received surveys immediately after the intervention and at 1 to 4 weeks after the post-intervention. Thematic analyses of qualitative feedback were performed using Braun and Clarke's thematic analysis framework. RESULTS: Inductive analyses revealed three main themes: (1) Developing self-acceptance through emotional reflection, (2) Changing expectations with neurosocial knowledge, and (3) Bonding and vulnerability in social interactions; all concepts intentionally targeted by the SBPI. Participants varied in their support of a guideline to exclude personal discussion of ED-related cognitions and behaviors in the group. CONCLUSIONS: As a whole, patients valued the combination of psychosocial education with group experientials focused on social behavior. Positive feedback from the SBPI suggests that adjunctive treatments that target mental-wellness constructs indirectly related to ED pathology may be helpful by allowing patients to see themselves as separable from the illness. Trial registration ClinicalTrials.gov, NCT0487758. Registered 7 May 2021-Retrospectively registered. https://clinicaltrials.gov/ct2/show/NCT04877158 .

Adult patients with eating disorders frequently report challenges in developing and maintaining supportive social relationships. A group therapy intervention utilizing structured art tasks in conjunction with psychoeducation about interpersonal relationships was piloted in twenty-four women with eating disorders. Participants provided written feedback about their experiences in the intervention. These responses were coded by three raters to identify consistent themes related to those experiences. Participants reported benefits related to self-acceptance and emotional regulation, valued learning about their brain and behaviors, and appreciated peer interactions in this setting. In sum, this adjunctive and structured outpatient group targeting social interactions was acceptable to outpatients with eating disorders.

Pilot data from the Self-Blame and Perspective-Taking Intervention for eating disorders.

Eating disorders (EDs) are characterized by altered eating behaviors and valuation of self-image, as well as difficulty establishing supportive social relationships. This pilot study evaluated feasibility, acceptability, and clinical responses to a novel and brief group-therapy intervention for EDs, the Self-Blame and Perspective-Taking Intervention (SBPI). The SBPI consisted of four sessions of experiential art therapy activities in conjunction with psychoeducation targeting interpersonal attributions and mentalization. Twenty-four outpatient, treatment-seeking women with EDs participated in the SBPI, with 87.5% completing the intervention and 94% rating their participation positively. ED symptoms, depression, anxiety, self-attribution bias, and self-esteem were assessed before (T1) and after participation (N = 20 at T2; N = 18 at T3). Separate repeated measures MANOVAs were performed to assess these clinical and self-concept variables. Relative to baseline, participants demonstrated significant improvements in two all self-concept measures: self-attribution bias, trait self-esteem and state self-esteem at T2. ED, depression, and anxiety symptoms were significantly decreased at both T2 (1-4 weeks post) and T3 (3-5 months post). The SBPI altered self-concept targets acutely and led to sustained clinical improvements. Future work is needed to evaluate how self-concept and social constructs are related to clinical symptom expression in EDs.

clc-2c is regulated by salinity, prolactin and extracellular osmolality in tilapia gill.

Teleosts inhabiting fresh water (FW) depend upon ion-absorptive ionocytes to counteract diffusive ion losses to the external environment. A Clc Cl- channel family member, Clc-2c, was identified as a conduit for basolateral Cl- transport by Na+/Cl- cotransporter 2 (Ncc2)-expressing ionocytes in stenohaline zebrafish (Danio rerio). It is unresolved whether Clc-2c/clc-2c is expressed in euryhaline species and how extrinsic and/or intrinsic factors modulate branchial clc-2c mRNA. Here, we investigated whether environmental salinity, prolactin (Prl) and osmotic conditions modulate clc-2c expression in euryhaline Mozambique tilapia (Oreochromis mossambicus). Branchial clc-2c and ncc2 mRNAs were enhanced in tilapia transferred from seawater (SW) to FW, whereas both mRNAs were attenuated upon transfer from FW to SW. Next, we injected hypophysectomized tilapia with ovine prolactin (oPrl) and observed a marked increase in clc-2c from saline-injected controls. To determine whether Prl regulates clc-2c in a gill-autonomous fashion, we incubated gill filaments in the presence of homologous tilapia Prls (tPrl177 and tPrl188). By 24 h, tPrl188 stimulated clc-2c expression ~5-fold from controls. Finally, filaments incubated in media ranging from 280 to 450 mosmol/kg for 3 and 6 h revealed that extracellular osmolality exerts a local effect on clc-2c expression; clc-2c was diminished by hyperosmotic conditions (450 mosmol/kg) compared with isosmotic controls (330 mosmol/kg). Our collective results suggest that hormonal and osmotic control of branchial clc-2c contributes to the FW adaptability of Mozambique tilapia. Moreover, we identify for the first time a regulatory link between Prl and a Clc Cl- channel in a vertebrate.

Hormonal regulation of aquaporin 3: opposing actions of prolactin and cortisol in tilapia gill.

Aquaporins (Aqps) are expressed within key osmoregulatory tissues where they mediate the movement of water and selected solutes across cell membranes. We leveraged the functional plasticity of Mozambique tilapia (Oreochromis mossambicus) gill epithelium to examine how Aqp3, an aquaglyceroporin, is regulated in response to osmoregulatory demands. Particular attention was paid to the actions of critical osmoregulatory hormones, namely, prolactin (Prl), growth hormone and cortisol. Branchial aqp3 mRNA levels were modulated following changes in environmental salinity, with enhanced aqp3 mRNA expression upon transfer from seawater to freshwater (FW). Accordingly, extensive Aqp3 immunoreactivity was localized to cell membranes of branchial epithelium in FW-acclimated animals. Upon transferring hypophysectomized tilapia to FW, we identified that a pituitary factor(s) is required for Aqp3 expression in FW. Replacement with ovine Prl (oPrl) was sufficient to stimulate Aqp3 expression in hypophysectomized animals held in FW, an effect blocked by coinjection with cortisol. Both oPrl and native tilapia Prls (tPrl177 and tPrl188) stimulated aqp3 in incubated gill filaments in a concentration-related manner. Consistent with in vivo responses, coincubation with cortisol blocked oPrl-stimulated aqp3 expression in vitro Our data indicate that Prl and cortisol act directly upon branchial epithelium to regulate Aqp3 in tilapia. Thus, within the context of the diverse actions of Prl on hydromineral balance in vertebrates, we define a new role for Prl as a regulator of Aqp expression.

Within-state availability of transition-to-adulthood services for youths with serious mental health conditions.

OBJECTIVE: This study describes the existence and nature of services within state child and adult mental health systems that support the transition from adolescence to adulthood. METHODS: State child and adult mental health administrators from all but one state were interviewed by telephone with a semistructured questionnaire regarding transition services in their state mental health system, such as supported housing, vocational support, preparation for independent living, and dual diagnosis treatment. Eight states were deemed sufficiently decentralized to render state-level administrator reports invalid. Specific service data from the remaining 41 states and the District of Columbia were analyzed with descriptive statistics. RESULTS: One-quarter of child state mental health systems and one-half of adult state mental health systems offered no transition services, and few provided any kind of transition service at more than one site. Most types of transition services were available at all in less than 20 percent of the states. CONCLUSIONS: Across the United States transition support services are lacking. The adult system in particular will require major transformation to provide the service capacity that is needed to meet the current standards of transition service accessibility for young Americans with serious mental health conditions.