SAF-A/hnRNP U binds polyphosphoinositides via a lysine rich polybasic motif located in the SAP domain

Polyphosphoinositides (PPIn) play essential functions as lipid signalling molecules and many of their functions have been elucidated in the cytoplasm. However, PPIn are also intranuclear where they contribute to chromatin remodelling, transcription and mRNA splicing. Using quantitative interactomics, we have previously identified PPIn-interacting proteins with roles in RNA processing/splicing including the heterogeneous nuclear ribonucleoprotein U (hnRNPU/SAF-A). In this study, hnRNPU was validated as a direct PPIn-interacting protein via 2 regions located in the N and C termini. Furthermore, deletion of the polybasic motif region located at aa 9-24 in its DNA binding SAP domain prevented PPIn interaction. In conclusion, these results are consistent with hnRNPU harbouring a polybasic region with dual functions in DNA and PPIn interaction.publishedVersio

DNA Topoisomerase IIα contributes to the early steps of adipogenesis in 3T3-L1 cells

DNA topoisomerases (Topo) are multifunctional enzymes resolving DNA topological problems such as those arising during DNA replication, transcription and mitosis. Mammalian cells express 2 class II isoforms, Topoisomerases IIα (Topo IIα) and IIβ (Topo IIβ), which have similar enzymatic properties but are differently expressed, in dividing and pluripotent cells, and in post-mitotic and differentiated cells respectively. Pre-adipocytes re-enter the cell cycle prior to committing to their differentiation and we hypothesised that Topo II could contribute to these processes. We show that Topo IIα expression in 3T3-L1 cells is induced within 16 h after the initiation of the differentiation programme, peaks at 24 h and rapidly declines thereafter. In contrast Topo IIβ was present both in pre-adipocytes and throughout differentiation. Inhibition of PI3K with LY294002, known to prevent adipocyte differentiation, consistently reduced the expression of Topo IIα, whereas a clear effect on Topo IIβ was not apparent. In addition, inhibition of mTOR with rapamycin also reduced the protein levels of Topo IIα. Using specific class IA PI3K catalytic subunit inhibitors, we show that p110α inhibition with A66 has the greatest reduction of Topo IIα expression and of differentiation, as measured by triglyceride storage. The timing of Topo IIα expression coincides with the mitotic clonal expansion (MCE) phase of differentiation and inhibition of Topo II with ICRF-187 during this stage decreased PPARγ1 and 2 protein levels and triglyceride storage, whereas inhibition later on has little impact. Moreover, the addition of ICRF-187 had no effect on the incorporation of EdU during S-phase at day 1 but lowered the relative cell numbers on day 2. ICRF-187 also induced an increase in the centri/pericentromeric heterochromatin localisation of Topo IIα, indicating a role for Topo IIα at these locations during MCE. In summary, we present evidence that Topo IIα plays an important role in adipogenesis during MCE and in a PI3K/mTOR-dependent manner. Considering that Topoisomerases II are targets in cancer chemotherapy, our results highlight that treatment of cancer with Topo II inhibitors may alter metabolic processes in the adipose tissue.publishedVersio

Contrasting the phospholipid profiles of two neoplastic cell lines reveal a high PC:PE ratio for SH-SY5Y cells relative to A431 cells

Lipids have been implicated in Parkinson's Disease (PD). We therefore studied the lipid profile of the neuroblastoma SH-SY5Y cell line, which is used extensively in PD research and compared it to that of the A431 epithelial cancer cell line. We have isolated whole cell extracts (WC) and plasma membrane (PM) fractions of both cell lines. The isolates were analyzed with 31P NMR. We observed a significant higher abundance of phosphatidylcholine (PC) for SH-SY5Y cells for both WC (55 ± 4.1%) and PM (63.3 ± 3.1%) compared to WC (40.5 ± 2.2%) and PM (43.4 ± 1.3%) of A431. Moreover, a higher abundance of phosphatidylethanolamine was detected for the WC of A431 compared to the SH-SY5Y. Using LC-MS/MS, we also determined the relative abundance of fatty acid (FA) moieties for each phospholipid class, finding that SH-SY5Y had high polyunsaturated FA levels, including arachidonic acid compared to A431 cells. When comparing our results to reported compositions of brain and neural tissues, we note the much higher PC levels, as well as very low levels of docosahexaenoic acid. However, relative levels of arachidonic acid and other polyunsaturated fatty acids were elevated, in line with what is desirable for a neural model system

Active Referral Intervention following Fragility Fractures Leads to Enhanced Osteoporosis Follow-Up Care

At one major urban academic medical center, patients aged 50 years and older with fragility fractures were identified and scheduled or assisted in referral into osteoporosis medical management appointments. We evaluated the efficacy of an active intervention program at overcoming the logistical barriers and improving proper osteoporosis follow-up for persons who have sustained a fragility fracture. Of 681 patients treated for defined fractures, 168 were eligible and consented for the study of fragility fractures. Of those enrolled, 91 (54.2%) had appropriate osteoporosis follow-up on initial interview, and overall 120 (71.4%) had successful osteoporosis follow-up following our active intervention. Seventy patients (41.7%) were deemed to have no osteoporosis follow-up, and, of these, 48 were successfully referred to a scheduling coordinator. The scheduling coordinator was able to contact 37 (77%) patients to schedule proper follow-up, and, of these, 29 (78.4%) confirmed receiving an appropriate follow-up appointment. Active intervention and assisted scheduling for patients with recent fragility fractures improved the self-reported rate of osteoporosis follow-up from 54.2% to 71.4%

HST/WFC3 Light Curve Confirms the Closest Exoplanet to Transit an M Dwarf is Terrestrial

Previous studies of the exoplanet LTT 1445Ac concluded that the light curve from the Transiting Exoplanet Survey Satellite (TESS) was consistent with both grazing and non-grazing geometries. As a result, the radius and hence density of the planet remained unknown. To resolve this ambiguity, we observed the LTT 1445 system for six spacecraft orbits of the Hubble Space Telescope (HST) using WFC3/UVIS imaging in spatial scan mode, including one partial transit of LTT 1445Ac. This imaging produces resolved light curves of each of the three stars in the LTT 1445 system. We confirm that the planet transits LTT 1445A and that LTT 1445C is the source of the rotational modulation seen in the TESS light curve, and we refine the estimate of the dilution factor for the TESS data. We perform a joint fit to the TESS and HST observations, finding that the transit of LTT 1445Ac is not grazing with 97% confidence. We measure a planetary radius of 1.10$_{-0.07}^{+0.10}$ R$_\oplus$. Combined with previous radial velocity observations, our analysis yields a planetary mass of $1.36\pm0.19$ M$_\oplus$ and a planetary density of 5.6$_{-1.5}^{+1.7}$ g cm$^{-3}$. LTT 1445Ac is an Earth analog with respect to its mass and radius, albeit with a higher instellation, and is therefore an exciting target for future atmospheric studies.Comment: Submitted to AJ. 9 pages, 7 figures, 3 table

Endometrial cancer cells exhibit high expression of p110beta and its selective inhibition induces variable responses on PI3K signaling, cell survival and proliferation

PTEN loss and constitutive activation of the class I phosphoinositide 3-kinase (PI3K) pathway are key drivers of endometrial tumorigenesis. In some cancer types, PTEN-deficient tumors are reliant on class I PI3K p110β (encoded by PIK3CB) activity but little is known about this contribution in endometrial tumorigenesis. In this study, we find that p110β is overexpressed in a panel of 7 endometrial cancer cell lines compared to non-transformed cells. Furthermore, in 234 clinically annotated patient samples, PIK3CB mRNA levels increase significantly in the early phase of tumorigenesis from precursors to low grade primary malignant lesions whereas PIK3CA levels are higher in non-endometrioid compared to endometrioid primary tumors. While high levels of either PIK3CA or PIK3CB associate with poor prognosis, only elevated PIK3CB mRNA levels correlate with a high cell cycle signature score in clinical samples. In cancer cell lines, p110α inhibition reduces cell viability by inducing cell death in PIK3CA mutant cells while p110β inhibition delayed proliferation in PTEN-deficient cells, but not in WT cells. Taken together, our findings suggest that PIK3CB/p110β contributes to some of the pleiotropic functions of PI3K in endometrial cancer, particularly in the early steps by contributing to cell proliferation

Clinical Study Active Referral Intervention following Fragility Fractures Leads to Enhanced Osteoporosis Follow-Up Care

At one major urban academic medical center, patients aged 50 years and older with fragility fractures were identified and scheduled or assisted in referral into osteoporosis medical management appointments. We evaluated the efficacy of an active intervention program at overcoming the logistical barriers and improving proper osteoporosis follow-up for persons who have sustained a fragility fracture. Of 681 patients treated for defined fractures, 168 were eligible and consented for the study of fragility fractures. Of those enrolled, 91 (54.2%) had appropriate osteoporosis follow-up on initial interview, and overall 120 (71.4%) had successful osteoporosis follow-up following our active intervention. Seventy patients (41.7%) were deemed to have no osteoporosis follow-up, and, of these, 48 were successfully referred to a scheduling coordinator. The scheduling coordinator was able to contact 37 (77%) patients to schedule proper follow-up, and, of these, 29 (78.4%) confirmed receiving an appropriate follow-up appointment. Active intervention and assisted scheduling for patients with recent fragility fractures improved the selfreported rate of osteoporosis follow-up from 54.2% to 71.4%

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

Polyphosphoinositides in the nucleus: Roadmap of their effectors and mechanisms of interaction

Biomolecular interactions between proteins and polyphosphoinositides (PPIn) are essential in the regulation of the vast majority of cellular processes. Consequently, alteration of these interactions is implicated in the development of many diseases. PPIn are phosphorylated derivatives of phosphatidylinositol and consist of seven species with different phosphate combinations. PPIn signal by recruiting proteins via canonical domains or short polybasic motifs. Although their actions are predominantly documented on cytoplasmic membranes, six of the seven PPIn are present within the nucleus together with the PPIn kinases, phosphatases and phospholipases that regulate their turnover. Importantly, the contribution of nuclear PPIn in the regulation of nuclear processes has led to an increased recognition of their importance compared to their more accepted cytoplasmic roles. This review summarises our knowledge on the identification and functional characterisation of nuclear PPIn-effector proteins as well as their mode of interactions, which tend to favour polybasic motifs.publishedVersionpublishedVersio

A polybasic motif in ErbB3-binding protein 1 (EBP1) has key functions in nucleolar localization and polyphosphoinositide interaction

Polyphosphoinositides (PPIns) are present in the nucleus where they participate in crucial nuclear processes, such as chromatin remodelling, transcription and mRNA processing. In a previous interactomics study, aimed to gain further insight into nuclear PPIns functions, we identified ErbB3 binding protein 1 (EBP1) as a potential nuclear PPIn-binding protein in a lipid pull-down screen. EBP1 is a ubiquitous and conserved protein, located in both the cytoplasm and nucleolus, and associated with cell proliferation and survival. In the present study, we show that EBP1 binds directly to several PPIns via two distinct PPIn-binding sites consisting of clusters of lysine residues and positioned at the N- and C-termini of the protein. Using interaction mutants, we show that the C-terminal PPIn-binding motif contributes the most to the localization of EBP1 in the nucleolus. Importantly, a K372N point mutation, located within the C-terminal motif and found in endometrial tumours, is sufficient to alter the nucleolar targeting of EBP1. Our study reveals also the presence of the class I phosphoinositide 3-kinase (PI3K) catalytic subunit p110β and its product PtdIns(3,4,5)P3 together with EBP1 in the nucleolus. Using NMR, we further demonstrate an association between EBP1 and PtdIns(3,4,5)P3 via both electrostatic and hydrophobic interactions. Taken together, these results show that EBP1 interacts directly with PPIns and associate with PtdIns(3,4,5)P3 in the nucleolus. The presence of p110β and PtdIns(3,4,5)P3 in the nucleolus indicates their potential role in regulating nucleolar processes, at least via EBP1