A pivotal mediator of actin dynamics is the protein cofilin, which promotes filament severing and depolymerization, facilitating the breakdown of existing filaments, and the enhancement of filament growth from newly created barbed ends. It does so in concert with actin interacting protein 1 (Aip1), which serves to accelerate cofilin's activity. While progress has been made in understanding its biochemical functions, the physiologic processes the cofilin/Aip1 complex regulates, particularly in higher organisms, are yet to be determined. We have generated an allelic series for WD40 repeat protein 1 (Wdr1), the mammalian homolog of Aip1, and report that reductions in Wdr1 function produce a dramatic phenotype gradient. While severe loss of function at the Wdr1 locus causes embryonic lethality, macrothrombocytopenia and autoinflammatory disease develop in mice carrying hypomorphic alleles. Macrothrombocytopenia is the result of megakaryocyte maturation defects, which lead to a failure of normal platelet shedding. Autoinflammatory disease, which is bone marrow-derived yet nonlymphoid in origin, is characterized by a massive infiltration of neutrophils into inflammatory lesions. Cytoskeletal responses are impaired in Wdr1 mutant neutrophils. These studies establish an essential requirement for Wdr1 in megakaryocytes and neutrophils, indicating that cofilin-mediated actin dynamics are critically important to the development and function of both cell types.

SummaryWhen migrating mesenchymal cells collide, they exhibit a 'contact inhibition of locomotion' response that results in reversal of their front-rear polarity by extension of a new leading edge, which enables their migration away from the opposing contacted cell. The critical cytoskeletal rearrangements underpinning these mutual repulsion events are currently unknown. We found that during fibroblast cell-cell collisions, microtubules at the region of contact increase their frequency of catastrophe, their rates of shrinkage and growth, and concomitantly, a new microtubule array is established at a new leading edge. We show that Rho and ROCK activity is necessary for this repulsion response, and we observed increased microtubule stabilisation as a consequence of ROCK inhibition. Importantly, partial destabilisation of microtubules, by co-treatment with a low dose of nocodazole, restored microtubule dynamics to that of untreated cells and rescued contact inhibition of locomotion in ROCK-inhibited cells. Although there was an increase in microtubule growth or shrinkage rates in Y27632 cell-cell collisions, these failed to reach the same level of dynamicity compared with untreated collisions. Our data suggest that microtubule dynamics at contact sites must increase beyond a threshold for a cell to switch its frontrear polarity, and that microtubule stabilisation can lead to a failure of contact inhibition of locomotion.

Abusing AAS is starting to become a public health concern that implies the need to implement educational programmes, which will educate and warn adolescents and mentors about the negative side effects of AAS abuse on the health of users.

Proteins of the actin depolymerizing factor (ADF)/cofilin family are ubiquitous among eukaryotes and are essential regulators of actin dynamics and function. Mammalian neurons express cofilin-1 as the major isoform, but ADF and cofilin-2 are also expressed. All isoforms bind preferentially and cooperatively along ADP-subunits in F-actin, affecting the filament helical rotation, and when either alone or when enhanced by other proteins, promotes filament severing and subunit turnover. Although self-regulating cofilin-mediated actin dynamics can drive motility without post-translational regulation, cells utilize many mechanisms to locally control cofilin, including cooperation/competition with other proteins. Newly identified post-translational modifications function with or are independent from the well-established phosphorylation of serine 3 and provide unexplored avenues for isoform specific regulation. Cofilin modulates actin transport and function in the nucleus as well as actin organization associated with mitochondrial fission and mitophagy. Under neuronal stress conditions, cofilin-saturated F-actin fragments can undergo oxidative cross-linking and bundle together to form cofilin-actin rods. Rods form in abundance within neurons around brain ischemic lesions and can be rapidly induced in neurites of most hippocampal and cortical neurons through energy depletion or glutamate-induced excitotoxicity. In ~20% of rodent hippocampal neurons, rods form more slowly in a receptor-mediated process triggered by factors intimately connected to disease-related dementias, e.g., amyloid-β in Alzheimer’s disease. This rod-inducing pathway requires a cellular prion protein, NADPH oxidase, and G-protein coupled receptors, e.g., CXCR4 and CCR5. Here, we will review many aspects of cofilin regulation and its contribution to synaptic loss and pathology of neurodegenerative diseases.

BackgroundADF/cofilin proteins are key modulators of actin dynamics in metastasis and invasion of cancer cells. Here we focused on the roles of ADF and cofilin-1 individually in the development of polarized migration of rat mammary adenocarcinoma (MTLn3) cells, which express nearly equal amounts of each protein. Small interference RNA (siRNA) technology was used to knockdown (KD) the expression of ADF and cofilin-1 independently.ResultsEither ADF KD or cofilin KD caused cell elongation, a reduction in cell area, a decreased ability to form invadopodia, and a decreased percentage of polarized cells after 180 s of epidermal growth factor stimulation. Moreover, ADF KD or cofilin KD increased the rate of cell migration and the time of lamellipodia protrusion but through different mechanisms: lamellipodia protrude more frequently in ADF KD cells and are more persistent in cofilin KD cells. ADF KD cells showed a significant increase in F-actin aggregates, whereas cofilin KD cells showed a significant increase in prominent F-actin bundles and increased cell adhesion. Focal adhesion area and cell adhesion in cofilin KD cells were returned to control levels by expressing exogenous cofilin but not ADF. Return to control rates of cell migration in ADF KD cells was achieved by expression of exogenous ADF but not cofilin, whereas in cofilin KD cells, expression of cofilin efficiently rescued control migration rates.ConclusionAlthough ADF and cofilin have many redundant functions, each of these isoforms has functional differences that affect F-actin structures, cell adhesion and lamellipodial dynamics, all of which are important determinants of cell migration.

2DG causes cytotoxicity in cancer cells by disrupting thiol metabolism while Doxorubicin (DOX) induces cytotoxicity in tumor cells by generating reactive oxygen species (ROS). Here we examined the combined cytotoxic action of 2DG and DOX in rapidly dividing T47D breast cancer cells vs. slowly growing MCF-7 breast cancer cells. T47D cells exposed to the combination of 2DG/DOX significantly decreased cell survival compared to controls, while 2DG/DOX had no effect on MCF-7 cells. 2DG/DOX also disrupted the oxidant status of T47D treated cells, decreased intracellular total glutathione and increased glutathione disulfide (%GSSG) compared to MCF-7 cells. Lipid peroxidation increased in T47D cells treated with 2DG and/or DOX, but not in MCF-7 cells. T47D cells were significantly protected by NAC, indicating that the combined treatment exerts its action by increasing ROS production and disrupting antioxidant stores. When we inhibited glutathione synthesis with BSO, T47D cells became more sensitive to 2DG/DOX-induced cytotoxicity, but NAC significantly reversed this cytotoxic effect. Finally, 2DG/DOX, and BSO significantly increased the %GSSG in T47D cells, an effect which was also reversed by NAC. Our results suggest that exposure of rapidly dividing breast cancer cells to 2DG/DOX enhances cytotoxicity via oxidative stress and via disruptions to thiol metabolism.

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BackgroundCord blood transplant is an accepted treatment for many malignant and non-malignant diseases. We sought to determine the feasibility of collecting cord blood in Jordan and the effect of maternal and fetal factors on the quality of the cord blood units.MethodsA total of 124 cord blood units were collected, and 75 (60%) cord blood units were included in this analysis. Cord blood volume, total nucleated cell (TNC) count, cell viability and CD34+ content were measured, and clonogenic assay was performed.ResultsThe mean volume of the collected units was 68.9 ml (range 40–115) with mean nucleated cell count of 6.5 x 108 (range 1–23.0). Our results showed a positive correlation between the volume of cord blood and TNC count (p=0.008), cell viability (p=0.001), CD34+ content (p=0.034) and the length of the umbilical cord (p=0.011). In addition, our results showed an inverse relation between the Colony Forming Unit-Granulocyte Macrophage (CFU-GM) concentration and the gestation duration (p=0.038).ConclusionWe conclude that it is feasible to collect cord blood units in Jordan with excellent TNC and CD34+ cell content. The volume of cord blood collected was associated with higher TNC count and CD34+ count. Efforts toward establishing public cord blood banks in our area are warranted.