Prolonged HPA axis dysregulation in postpartum depression associated with adverse early life experiences: A cross-species translational study.

Childhood and adolescent stress increase the risk of postpartum depression (PPD), often providing an increased probability of treatment refractoriness. Nevertheless, the mechanisms linking childhood/adolescent stress to PPD remain unclear. Our study investigated the longitudinal effects of adolescent stress on the hypothalamic-pituitary-adrenal (HPA) axis and postpartum behaviors in mice and humans. Adolescent social isolation prolonged glucocorticoid elevation, leading to long-lasting postpartum behavioral changes in female mice. These changes were unresponsive to current PPD treatments but improved with post-delivery glucocorticoid receptor antagonist treatment. Childhood/adolescent stress significantly impacted HPA axis dysregulation and PPD in human females. Repurposing glucocorticoid receptor antagonists for some cases of treatment-resistant PPD may be considered.

Mapping Ligand Interactions of Bromodomains BRD4 and ATAD2 with FragLites and PepLites─Halogenated Probes of Druglike and Peptide-like Molecular Interactions.

The development of ligands for biological targets is critically dependent on the identification of sites on proteins that bind molecules with high affinity. A set of compounds, called FragLites, can identify such sites, along with the interactions required to gain affinity, by X-ray crystallography. We demonstrate the utility of FragLites in mapping the binding sites of bromodomain proteins BRD4 and ATAD2 and demonstrate that FragLite mapping is comparable to a full fragment screen in identifying ligand binding sites and key interactions. We extend the FragLite set with analogous compounds derived from amino acids (termed PepLites) that mimic the interactions of peptides. The output of the FragLite maps is shown to enable the development of ligands with leadlike potency. This work establishes the use of FragLite and PepLite screening at an early stage in ligand discovery allowing the rapid assessment of tractability of protein targets and informing downstream hit-finding.

Discriminative SKP2 Interactions with CDK-Cyclin Complexes Support a Cyclin A-Specific Role in p27KIP1 Degradation.

The SCFSKP2 ubiquitin ligase relieves G1 checkpoint control of CDK-cyclin complexes by promoting p27KIP1 degradation. We describe reconstitution of stable complexes containing SKP1-SKP2 and CDK1-cyclin B or CDK2-cyclin A/E, mediated by the CDK regulatory subunit CKS1. We further show that a direct interaction between a SKP2 N-terminal motif and cyclin A can stabilize SKP1-SKP2-CDK2-cyclin A complexes in the absence of CKS1. We identify the SKP2 binding site on cyclin A and demonstrate the site is not present in cyclin B or cyclin E. This site is distinct from but overlapping with features that mediate binding of p27KIP1 and other G1 cyclin regulators to cyclin A. We propose that the capacity of SKP2 to engage with CDK2-cyclin A by more than one structural mechanism provides a way to fine tune the degradation of p27KIP1 and distinguishes cyclin A from other G1 cyclins to ensure orderly cell cycle progression.

FragLites-Minimal, Halogenated Fragments Displaying Pharmacophore Doublets. An Efficient Approach to Druggability Assessment and Hit Generation.

Identifying ligand binding sites on proteins is a critical step in target-based drug discovery. Current approaches to this require resource-intensive screening of large libraries of lead-like or fragment molecules. Here, we describe an efficient and effective experimental approach to mapping interaction sites using a set of halogenated compounds expressing paired hydrogen-bonding motifs, termed FragLites. The FragLites identify productive drug-like interactions, which are identified sensitively and unambiguously by X-ray crystallography, exploiting the anomalous scattering of the halogen substituent. This mapping of protein interaction surfaces provides an assessment of druggability and can identify efficient start points for the de novo design of hit molecules incorporating the interacting motifs. The approach is illustrated by mapping cyclin-dependent kinase 2, which successfully identifies orthosteric and allosteric sites. The hits were rapidly elaborated to develop efficient lead-like molecules. Hence, the approach provides a new method of identifying ligand sites, assessing tractability and discovering new leads.

Differences in the Conformational Energy Landscape of CDK1 and CDK2 Suggest a Mechanism for Achieving Selective CDK Inhibition.

Dysregulation of the cell cycle characterizes many cancer subtypes, providing a rationale for developing cyclin-dependent kinase (CDK) inhibitors. Potent CDK2 inhibitors might target certain cancers in which CCNE1 is amplified. However, current CDK2 inhibitors also inhibit CDK1, generating a toxicity liability. We have used biophysical measurements and X-ray crystallography to investigate the ATP-competitive inhibitor binding properties of cyclin-free and cyclin-bound CDK1 and CDK2. We show that these kinases can readily be distinguished by such inhibitors when cyclin-free, but not when cyclin-bound. The basis for this discrimination is unclear from either inspection or molecular dynamics simulation of ligand-bound CDKs, but is reflected in the contacts made between the kinase N- and C-lobes. We conclude that there is a subtle but profound difference between the conformational energy landscapes of cyclin-free CDK1 and CDK2. The unusual properties of CDK1 might be exploited to differentiate CDK1 from other CDKs in future cancer therapeutic design.

Structural insights into the functional diversity of the CDK-cyclin family.

Since their characterization as conserved modules that regulate progression through the eukaryotic cell cycle, cyclin-dependent protein kinases (CDKs) in higher eukaryotic cells are now also emerging as significant regulators of transcription, metabolism and cell differentiation. The cyclins, though originally characterized as CDK partners, also have CDK-independent roles that include the regulation of DNA damage repair and transcriptional programmes that direct cell differentiation, apoptosis and metabolic flux. This review compares the structures of the members of the CDK and cyclin families determined by X-ray crystallography, and considers what mechanistic insights they provide to guide functional studies and distinguish CDK- and cyclin-specific activities. Aberrant CDK activity is a hallmark of a number of diseases, and structural studies can provide important insights to identify novel routes to therapy.

Fitness Profiles of Elite Adolescent Irish Rugby Union Players.

Wood, DJ, Coughlan, GF, and Delahunt, E. Fitness profiles of elite adolescent Irish rugby union players. J Strength Cond Res 32(1): 105-112, 2018-Rugby unions throughout the world are implementing player development models to prepare young players to meet the demands of professional rugby union. An example of this is the Irish Rugby Football Union Long Term Player Development model. The purpose of this study was to provide normative data relating to the physical fitness of elite adolescent Irish rugby union players and determine the differences in the physical capacities between players in the forward and back units as well as to provide descriptive data for the position categorizations within these units for this unique population. Players in the forward unit were significantly taller and heavier than players in the back unit (1.85 ± 0.06 m and 96.88 ± 9.00 kg vs. 1.79 ± 0.05 m and 81.97 ± 7.09 kg, respectively). Forwards (38.37 ± 4.00 cm) had a significantly lower countermovement jump height than backs (41.31 ± 4.44 cm). Forwards had a significantly lower triple hop for the distance score on their right (5.78 ± 0.52 m) and left (5.78 ± 0.55 m) legs compared with backs (6.26 ± 0.42 m and 6.33 ± 0.45 m, respectively). Forwards (1.85 ± 0.07 seconds) had a significantly higher 10-m sprint time than backs (1.77 ± 0.06 seconds). Furthermore, forwards (675.90 ± 82.46 m) had a significantly lower 150-m shuttle test score than backs (711.71 ± 27.46 m). The results of this study provide normative data for players who currently possess underage international potential and could be used by strength and conditioning coaches to guide the selection of players through talent identification processes.

Tibial loading increases osteogenic gene expression and cortical bone volume in mature and middle-aged mice.

There are conflicting data on whether age reduces the response of the skeleton to mechanical stimuli. We examined this question in female BALB/c mice of different ages, ranging from young to middle-aged (2, 4, 7, 12 months). We first assessed markers of bone turnover in control (non-loaded) mice. Serum osteocalcin and CTX declined significantly from 2 to 4 months (p<0.001). There were similar age-related declines in tibial mRNA expression of osteoblast- and osteoclast-related genes, most notably in late osteoblast/matrix genes. For example, Col1a1 expression declined 90% from 2 to 7 months (p<0.001). We then assessed tibial responses to mechanical loading using age-specific forces to produce similar peak strains (-1300 µÎµ endocortical; -2350 µÎµ periosteal). Axial tibial compression was applied to the right leg for 60 cycles/day on alternate days for 1 or 6 weeks. qPCR after 1 week revealed no effect of loading in young (2-month) mice, but significant increases in osteoblast/matrix genes in older mice. For example, in 12-month old mice Col1a1 was increased 6-fold in loaded tibias vs. controls (p = 0.001). In vivo microCT after 6 weeks revealed that loaded tibias in each age group had greater cortical bone volume (BV) than contralateral control tibias (p<0.05), due to relative periosteal expansion. The loading-induced increase in cortical BV was greatest in 4-month old mice (+13%; p<0.05 vs. other ages). In summary, non-loaded female BALB/c mice exhibit an age-related decline in measures related to bone formation. Yet when subjected to tibial compression, mice from 2-12 months have an increase in cortical bone volume. Older mice respond with an upregulation of osteoblast/matrix genes, which increase to levels comparable to young mice. We conclude that mechanical loading of the tibia is anabolic for cortical bone in young and middle-aged female BALB/c mice.

Ursodeoxycholic acid modulates histone acetylation and induces differentiation and senescence.

Agents that can modulate colonic environment and control dysregulated signaling are being evaluated for their chemopreventive potential in colon cancer. Ursodeoxycholate (UDCA) has shown chemopreventive potential in preclinical and animal models of colon cancer, but the mechanism behind it remains unknown. Here biological effects of UDCA were examined to understand mechanism behind its chemoprevention in colon cancer. Our data suggests that UDCA can suppress growth in a wide variety of cancer cell lines and can induce low level of apoptosis in colon cancer cells. We also found that UDCA treatment induces alteration in morphology, increased cell size, upregulation of cytokeratin 8, 18 and 19 and E-cadherin, cytokeratin remodeling and accumulation of lipid droplets, suggesting that UDCA induces differentiation in colon carcinoma cells. Our results also suggest significant differences in UDCA and sodium butyrate induced functional differentiation. We also report for the first time that UDCA can induce senescence in colon cancer cells as assessed by flattened, spread out and vacuolated morphology as well as by senescence marker beta-galactosidase staining. We also found that UDCA inhibits the telomerase activity. Surprisingly, we found that UDCA is not a histone deacytylase inhibitor but instead induces hypoacetylation of histones unlike hyperacetylation induced by sodium butyrate. Our results also suggest that, although UDCA induced senescence is p53, p21 and Rb independent, HDAC6 appears to be important in UDCA induced senescence. In summary, our data shows that UDCA modulates chromatin by inducing histone hypoacetylation and induces differentiation and senescence in colon cancer cells.