Preventing Sexual Harassment in Higher Education: A Framework for Prevention Science Program Development.

Sexual harassment is an intractable problem that harms the students, community, culture, and success of institutes of higher education (IHEs). The alarming prevalence of sexual harassment at IHEs highlights the urgent need for effective prevention programs. However, there are few empirically supported preventive interventions that effectively target the factors that most impact the determinants, trajectory, and short- and intermediate-term effects of sexual harassment. In this paper, we overview the problem of sexual harassment and propose an organizing framework to help IHEs develop effective interventions to prevent sexual harassment. Guided by prevention science, we propose a framework-modified from SAMHSA's (2019) guidelines for prevention practitioners-that underscores the criticality of trauma- and equity-informed characteristics in prevention programs. We offer a discussion on how IHEs must consider and evaluate the empirical evidence of effectiveness, flexibility, cultural competency, and sustainability when developing and adapting prevention programs to reduce and-ultimately-ameliorate sexual harassment. We conclude with recommendations that can provide a roadmap for higher education stakeholders and researchers to prevent this urgent public health concern.

The multidimensional role of the Wnt/ß-catenin signaling pathway in human malignancies.

Several signaling pathways have been identified as important for developmental processes. One of such important cascades is the Wnt/ß-catenin signaling pathway, which can regulate various physiological processes such as embryonic development, tissue homeostasis, and tissue regeneration; while its dysregulation is implicated in several pathological conditions especially cancers. Interestingly, deregulation of the Wnt/ß-catenin pathway has been reported to be closely associated with initiation, progression, metastasis, maintenance of cancer stem cells, and drug resistance in human malignancies. Moreover, several genetic and experimental models support the inhibition of the Wnt/ß-catenin pathway to answer the key issues related to cancer development. The present review focuses on different regulators of Wnt pathway and how distinct mutations, deletion, and amplification in these regulators could possibly play an essential role in the development of several cancers such as colorectal, melanoma, breast, lung, and leukemia. Additionally, we also provide insights on diverse classes of inhibitors of the Wnt/ß-catenin pathway, which are currently in preclinical and clinical trial against different cancers.

Interphase Chromosomes in Replicative Senescence: Chromosome Positioning as a Senescence Biomarker and the Lack of Nuclear Motor-Driven Chromosome Repositioning in Senescent Cells.

This study demonstrates, and confirms, that chromosome territory positioning is altered in primary senescent human dermal fibroblasts (HDFs). The chromosome territory positioning pattern is very similar to that found in HDFs made quiescent either by serum starvation or confluence; but not completely. A few chromosomes are found in different locations. One chromosome in particular stands out, chromosome 10, which is located in an intermediate location in young proliferating HDFs, but is found at the nuclear periphery in quiescent cells and in an opposing location of the nuclear interior in senescent HDFs. We have previously demonstrated that individual chromosome territories can be actively and rapidly relocated, with 15 min, after removal of serum from the culture media. These chromosome relocations require nuclear motor activity through the presence of nuclear myosin 1ß (NM1ß). We now also demonstrate rapid chromosome movement in HDFs after heat-shock at 42°C. Others have shown that heat shock genes are actively relocated using nuclear motor protein activity via actin or NM1ß (Khanna et al., 2014; Pradhan et al., 2020). However, this current study reveals, that in senescent HDFs, chromosomes can no longer be relocated to expected nuclear locations upon these two types of stimuli. This coincides with a entirely different organisation and distribution of NM1ß within senescent HDFs.

Chromosome territory relocation paradigm during DNA damage response: Some insights from molecular biology to physics.

Among the many facets of DNA damage response (DDR), relocation of chromosome territories (CTs) is most intriguing. We have previously reported that cisplatin induced DDR in human dermal fibroblasts led to relocation of CTs 12, 15 from the nuclear periphery to its interior while CTs 19, 17 repositioned from the interior to its periphery. Studies of CT relocation remain nascent as we begin unraveling the role of key players in DDR to demonstrate its mechanistic basis. Consolidating our recent reports, we argue that γH2AX-signaling leads to enhanced recruitment of nuclear myosin 1 (NM1) to chromatin, which via its motor function, results in CT repositioning. Next, we invoke a novel systems-level theory that subsumed CTs as pairs, not solo entities, to present the physical basis for plasticity in interphase CT arrangement. Subsequently, we posited that our systems-level theory describes a unified physical basis for non-random positioning of CTs in interphase nuclei across disparate eukaryotes.

Systems-level chromosomal parameters represent a suprachromosomal basis for the non-random chromosomal arrangement in human interphase nuclei.

Forty-six chromosome territories (CTs) are positioned uniquely in human interphase nuclei, wherein each of their positions can range from the centre of the nucleus to its periphery. A non-empirical basis for their non-random arrangement remains unreported. Here, we derive a suprachromosomal basis of that overall arrangement (which we refer to as a CT constellation), and report a hierarchical nature of the same. Using matrix algebra, we unify intrinsic chromosomal parameters (e.g., chromosomal length, gene density, the number of genes per chromosome), to derive an extrinsic effective gene density matrix, the hierarchy of which is dominated largely by extrinsic mathematical coupling of HSA19, followed by HSA17 (human chromosome 19 and 17, both preferentially interior CTs) with all CTs. We corroborate predicted constellations and effective gene density hierarchy with published reports from fluorescent in situ hybridization based microscopy and Hi-C techniques, and delineate analogous hierarchy in disparate vertebrates. Our theory accurately predicts CTs localised to the nuclear interior, which interestingly share conserved synteny with HSA19 and/or HSA17. Finally, the effective gene density hierarchy dictates how permutations among CT position represents the plasticity within its constellations, based on which we suggest that a differential mix of coding with noncoding genome modulates the same.

Chromosome territory relocation during DNA repair requires nuclear myosin 1 recruitment to chromatin mediated by ϒ-H2AX signaling.

During DNA damage response (DDR), certain gene rich chromosome territories (CTs) relocate to newer positions within interphase nuclei and revert to their native locations following repair. Such dynamic relocation of CTs has been observed under various cellular conditions, however, the underlying mechanistic basis of the same has remained largely elusive. In this study, we aim to understand the temporal and molecular details of such crosstalk between DDR signaling and CT relocation dynamics. We demonstrate that signaling at DNA double strand breaks (DSBs) by the phosphorylated histone variant (ϒ-H2AX) is a pre-requisite for damage induced CT relocation, as cells deficient in ϒ-H2AX signaling fail to exhibit such a response. Inhibition of Rad51 or DNA Ligase IV mediated late steps of double strand break repair does not seem to abrogate CT relocation completely. Upon DNA damage, an increase in the levels of chromatin bound motor protein nuclear myosin 1 (NM1) ensues, which appears to be functionally linked to ϒ-H2AX signaling. Importantly, the motor function of NM1 is essential for its recruitment to chromatin and CT relocation following damage. Taking these observations together, we propose that early DDR sensing and signaling result in NM1 recruitment to chromosomes which in turn guides DNA damage induced CT relocation.

The non-random repositioning of whole chromosomes and individual gene loci in interphase nuclei and its relevance in disease, infection, aging, and cancer.

The genomes of a wide range of different organisms are non-randomly organized within interphase nuclei. Chromosomes and genes can be moved rapidly, with direction, to new non-random locations within nuclei upon a stimulus such as a signal to initiate differentiation, quiescence or senescence, or also the application of heat or an infection with a pathogen. It is now becoming increasingly obvious that chromosome and gene position can be altered in diseases such as cancer and other syndromes that are affected by changes to nuclear architecture such as the laminopathies. This repositioning seems to affect gene expression in these cells and may play a role in progression of the disease. We have some evidence in breast cancer cells and in the premature aging disease Hutchinson-Gilford Progeria that an aberrant nuclear envelope may lead to genome repositioning and correction of these nuclear envelope defects can restore proper gene positioning and expression in both disease situations.Although spatial positioning of the genome probably does not entirely control expression of genes, it appears that spatio-epigenetics may enhance the control over gene expression globally and/or is deeply involved in regulating specific sets of genes. A deviation from normal spatial positioning of the genome for a particular cell type could lead to changes that affect the future health of the cell or even an individual.

Chromosome territories reposition during DNA damage-repair response.

BACKGROUND: Local higher-order chromatin structure, dynamics and composition of the DNA are known to determine double-strand break frequencies and the efficiency of repair. However, how DNA damage response affects the spatial organization of chromosome territories is still unexplored. RESULTS: Our report investigates the effect of DNA damage on the spatial organization of chromosome territories within interphase nuclei of human cells. We show that DNA damage induces a large-scale spatial repositioning of chromosome territories that are relatively gene dense. This response is dose dependent, and involves territories moving from the nuclear interior to the periphery and vice versa. Furthermore, we have found that chromosome territory repositioning is contingent upon double-strand break recognition and damage sensing. Importantly, our results suggest that this is a reversible process where, following repair, chromosome territories re-occupy positions similar to those in undamaged control cells. CONCLUSIONS: Thus, our report for the first time highlights DNA damage-dependent spatial reorganization of whole chromosomes, which might be an integral aspect of cellular damage response.

Farnesyltransferase inhibitor treatment restores chromosome territory positions and active chromosome dynamics in Hutchinson-Gilford progeria syndrome cells.

BACKGROUND: Hutchinson-Gilford progeria syndrome (HGPS) is a premature ageing syndrome that affects children leading to premature death, usually from heart infarction or strokes, making this syndrome similar to normative ageing. HGPS is commonly caused by a mutation in the A-type lamin gene, LMNA (G608G). This leads to the expression of an aberrant truncated lamin A protein, progerin. Progerin cannot be processed as wild-type pre-lamin A and remains farnesylated, leading to its aberrant behavior during interphase and mitosis. Farnesyltransferase inhibitors prevent the accumulation of farnesylated progerin, producing a less toxic protein. RESULTS: We have found that in proliferating fibroblasts derived from HGPS patients the nuclear location of interphase chromosomes differs from control proliferating cells and mimics that of control quiescent fibroblasts, with smaller chromosomes toward the nuclear interior and larger chromosomes toward the nuclear periphery. For this study we have treated HGPS fibroblasts with farnesyltransferase inhibitors and analyzed the nuclear location of individual chromosome territories. We have found that after exposure to farnesyltransferase inhibitors mis-localized chromosome territories were restored to a nuclear position akin to chromosomes in proliferating control cells. Furthermore, not only has this treatment afforded chromosomes to be repositioned but has also restored the machinery that controls their rapid movement upon serum removal. This machinery contains nuclear myosin 1ß, whose distribution is also restored after farnesyltransferase inhibitor treatment of HGPS cells. CONCLUSIONS: This study not only progresses the understanding of genome behavior in HGPS cells but demonstrates that interphase chromosome movement requires processed lamin A.

Rapid chromosome territory relocation by nuclear motor activity in response to serum removal in primary human fibroblasts.

BACKGROUND: Radial chromosome positioning in interphase nuclei is nonrandom and can alter according to developmental, differentiation, proliferation, or disease status. However, it is not yet clear when and how chromosome repositioning is elicited. RESULTS: By investigating the positioning of all human chromosomes in primary fibroblasts that have left the proliferative cell cycle, we have demonstrated that in cells made quiescent by reversible growth arrest, chromosome positioning is altered considerably. We found that with the removal of serum from the culture medium, chromosome repositioning took less than 15 minutes, required energy and was inhibited by drugs affecting the polymerization of myosin and actin. We also observed that when cells became quiescent, the nuclear distribution of nuclear myosin 1 beta was dramatically different from that in proliferating cells. If we suppressed the expression of nuclear myosin 1 beta by using RNA-interference procedures, the movement of chromosomes after 15 minutes in low serum was inhibited. When high serum was restored to the serum-starved cultures, chromosome repositioning was evident only after 24 to 36 hours, and this coincided with a return to a proliferating distribution of nuclear myosin 1 beta. CONCLUSIONS: These findings demonstrate that genome organization in interphase nuclei is altered considerably when cells leave the proliferative cell cycle and that repositioning of chromosomes relies on efficient functioning of an active nuclear motor complex that contains nuclear myosin 1 beta.

Progeria, the nucleolus and farnesyltransferase inhibitors.

HGPS (Hutchinson-Gilford progeria syndrome) is a rare genetic disease affecting children causing them to age and die prematurely. The disease is typically due to a point mutation in the coding sequence for the nuclear intermediate-type filament protein lamin A and gives rise to a dominant-negative splice variant named progerin. Accumulation of progerin within nuclei causes disruption to nuclear structure, causes and premature replicative senescence and increases apoptosis. Now it appears that accumulation of progerin may have more widespread effects than previously thought since the demonstration that the presence and distribution of some nucleolar proteins are also adversely affected in progeria cells. One of the major breakthroughs both in the lamin field and for this syndrome is that many of the cellular defects observed in HGPS patient cells and model systems can be restored after treatment with a class of compounds known as FTIs (farnesyltransferase inhibitors). Indeed, it is demonstrated that FTI-277 is able to completely restore nucleolar antigen localization in treated progeria cells. This is encouraging news for the HGPS patients who are currently undergoing clinical trials with FTI treatment.

Nuclear motors and nuclear structures containing A-type lamins and emerin: is there a functional link?

Rapid interphase chromosome territory repositioning appears to function through the action of nuclear myosin and actin, in a nuclear motor complex. We have found that chromosome repositioning when cells leave the cell cycle is not apparent in cells that have mutant lamin A or that are lacking emerin. We discuss the possibility that there is a functional intranuclear complex comprising four proteins: nuclear actin, lamin A, emerin and nuclear myosin. If any of the components are lacking or aberrant, then the nuclear motor complex involved in moving chromosomes or genes will be dysfunctional, leading to an inability to move chromosomes in response to signalling events.

Alterations to nuclear architecture and genome behavior in senescent cells.

The organization of the genome within interphase nuclei, and how it interacts with nuclear structures is important for the regulation of nuclear functions. Many of the studies researching the importance of genome organization and nuclear structure are performed in young, proliferating, and often transformed cells. These studies do not reveal anything about the nucleus or genome in nonproliferating cells, which may be relevant for the regulation of both proliferation and replicative senescence. Here, we provide an overview of what is known about the genome and nuclear structure in senescent cells. We review the evidence that nuclear structures, such as the nuclear lamina, nucleoli, the nuclear matrix, nuclear bodies (such as promyelocytic leukemia bodies), and nuclear morphology all become altered within growth-arrested or senescent cells. Specific alterations to the genome in senescent cells, as compared to young proliferating cells, are described, including aneuploidy, chromatin modifications, chromosome positioning, relocation of heterochromatin, and changes to telomeres.

Cystine/glutamate antiporter expression in retinal Müller glial cells: implications for DL-alpha-aminoadipate toxicity.

A cytotoxicity of glutamate or related amino acids (10 mM) mediated by a cystine/glutamate antiporter (system Xc) has recently been demonstrated in N18 neuroblastoma-rat retina hybrid (N18RE105) cells and C6 glioma cells. The antiporter usually transports glutamate outside and cystine inside, thereby maintaining cellular concentrations of glutathione. High concentrations of glutamate inhibit cystine uptake and lead to depletion of cellular levels of glutathione. Among related amino acids, DL-alpha-aminoadipic acid (DL-alpha-AAA), which is well known as a selective gliotoxin in the retina, is also toxic to these cells. However, this does not explain why DL-alpha-AAA acts gliospecifically on the retina. To answer this question we first examined the effects of DL-alpha-AAA on the [35S]cystine uptake with parental N18 neuroblastoma cells and rat retina of the hybrid cells. DL-alpha-AAA showed a competitive inhibition of [35S]cystine uptake in the rat retina but not in the N18 cells. Such a competitive inhibition of cystine uptake by DL-alpha-AAA could also be seen in the carp retina. The cystine uptake with carp retina was mainly Na(+)-independent and Cl(-)-dependent as already described as a characteristic ion dependency of the Xc antiporter. We next examined the effects of exogenous cystine on the glutamate release from the retina. Cystine (1 mM) actually induced a glutamate release approximately twice that of the control. Furthermore, the glutamate release induced by cystine was also Na(+)-independent and Cl(-)-dependent, and was blocked by DL-alpha-AAA. An autoradiogram of [35S]cystine uptake in the carp retina showed typical radial glial Müller cells. A large incorporation of [35S]cystine into retinal glutathione fraction was detected by a high pressure liquid chromatography method during a 1-4-h incubation. A significant or large decrease of retinal levels of glutathione was observed one day ater an intravitreal injection of 8 mumol DL-alpha-AAA or L-alpha-AAA, respectively. Buthionine sulfoximine (2.5 mumol), a specific inhibitor of glutathione synthesis, induced a large decrease of retinal levels of glutathione and a loss of electroretinographic b-wave 20-30 h after treatment. Taken together, our present data with rat and carp retinas strongly indicate that the expression of cystine/glutamate antiporter is enriched in the retina, particularly in the glial Müller cells which have a rapid turnover pool for glutathione. The gliotoxin DL-alpha-AAA inhibits cystine uptake through this antiporter on the glial cells and elicits reduction of cellular levels of glutathione.(ABSTRACT TRUNCATED AT 400 WORDS)

Dopamine release via protein kinase C activation in the fish retina.

Calcium-dependent phospholipid-sensitive protein kinase [protein kinase C (PKC)] was partially purified from the carp (Cyprinus carpio) retina through DE 52 ion exchange and Cellulofine gel filtration chromatography. The phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) activated PKC in the nanomolar range. A major 38-kDa protein in the retinal supernatants (105,000 g) was phosphorylated in vitro by PKC during a short period (3 min). Other phosphoproteins also appeared during a further prolonged period (greater than 15 min). Rod-bipolar and dopamine (DA) interplexiform cells in the fish retina were immunoreactive to a monoclonal antibody to PKC (alpha/beta-subtype). The PKC antibody recognized a 78-kDa native PKC enzyme by means of an immunoblotting method. Subsequently, the effects of two kinds of PKC activators were investigated on [3H]DA release from retinal cell fractions containing DA cells that had been preloaded with [3H]DA. A phorbol ester (TPA) induced a calcium- and dose-dependent [3H]DA release during a short period (2 min), with the minimal effective dose being approximately 1 nM. Other phorbols having no tumor-promoting activity, such as 4 beta-phorbol and 4 alpha-phorbol 12,13-didecanoate, were ineffective on [3H]DA release. A synthetic diacylglycerol [1-oleoyl-2-acetylglycerol (OAG)], which is an endogenous PKC activator, was also able to induce a significant release of [3H]DA. Furthermore, TPA was found to release endogenous DA from isolated fish retina by a highly sensitive HPLC with electrochemical detection method. The OAG- or TPA-induced [3H]DA or DA release was completely blocked by inhibitors of PKC, such as 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7) and staurosporine.(ABSTRACT TRUNCATED AT 250 WORDS)

GABAergic inhibition on dopamine cells of the fish retina: a [3H]dopamine release study with isolated fractions.

Inner retinal cells including dopamine (DA) cells were isolated and fractionated from the carp (Cyprinus carpio) retina by an enzyme cell dissociation and metrizamide gradient centrifugation method. When gamma-aminobutyric acid (GABA) antagonists (bicuculline and picrotoxin) were added into the perfusate over such a cell fraction, they stimulated the release of [3H]DA which had been preloaded in the cell fraction. The action of GABA antagonists was dose and Ca2+ dependent. Their minimal effective concentration was very low (0.5 microM). A similar action was elicited by high K+. In the presence of excess GABA, this stimulatory action of GABA antagonists and high K+ on [3H]DA release was completely abolished. To interpret the action of GABA antagonists on DA cells, isolated cell fractions were preincubated with GABAse. After such a treatment, the stimulatory effects of GABA antagonists and high K+ on [3H]DA release were differentiated from each other; the former disappeared whereas the latter remained unchanged. The data strongly suggest that GABA inhibits the DA release from retinal DA cells and thus the GABA antagonists affect [3H]DA release from cell fractions not by a direct membrane action but by a disinhibition mechanism via GABA receptors on the DA cell bodies.