Neural Wiskott-Aldrich syndrome protein (N-WASP) promotes distant metastasis in pancreatic ductal adenocarcinoma via activation of LOXL2.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive solid malignancies. A specific mechanism of its metastasis has not been established. In this study, we investigated whether Neural Wiskott-Aldrich syndrome protein (N-WASP) plays a role in distant metastasis of PDAC. We found that N-WASP is markedly expressed in clinical patients with PDAC. Clinical analysis showed a notably more distant metastatic pattern in the N-WASP-high group compared to the N-WASP-low group. N-WASP was noted to be a novel mediator of epithelial-mesenchymal transition (EMT) via gene expression profile studies. Knockdown of N-WASP in pancreatic cancer cells significantly inhibited cell invasion, migration, and EMT. We also observed positive association of lysyl oxidase-like 2 (LOXL2) and focal adhesion kinase (FAK) with the N-WASP-mediated response, wherein EMT and invadopodia function were modulated. Both N-WASP and LOXL2 depletion significantly reduced the incidence of liver and lung metastatic lesions in orthotopic mouse models of pancreatic cancer. These results elucidate a novel role for N-WASP signaling associated with LOXL2 in EMT and invadopodia function, with respect to regulation of intercellular communication in tumor cells for promoting pancreatic cancer metastasis. These findings may aid in the development of therapeutic strategies against pancreatic cancer.

Spectrum of WAS gene mutations in Vietnamese patients with Wiskott-Aldrich syndrome.

BACKGROUND: WAS gene mutational analysis is crucial to establish a definite diagnosis of Wiskott-Aldrich syndrome (WAS). Data on the genetic background of WAS in Vietnamese patients have not been reported. METHODS: We recruited 97 male, unrelated patients with WAS and analyzed WAS gene mutation using Sanger sequencing technology. RESULTS: We identified 36 distinct hemizygous pathogenic mutations, with 17 novel variants, from 38 patients in the entire cohort (39.2%). The mutational spectrum included 14 missense, 12 indel, five nonsense, four splicing, and one non-stop mutations. Most mutations appear only once, with the exception of c.37C>T (p.R13X) and c.374G>A (p.G125E) each of which occurs twice in unrelated patients. CONCLUSION: Our data enrich the mutational spectrum of the WAS gene and are crucial for understanding the genetic background of WAS and for supporting genetic counseling.

Both Las17-binding sites on Arp2/3 complex are important for branching nucleation and assembly of functional endocytic actin networks in S. cerevisiae.

Arp2/3 complex nucleates branched actin filaments that drive membrane invagination during endocytosis and leading-edge protrusion in lamellipodia. Arp2/3 complex is maximally activated in vitro by binding of a WASP family protein to two sites-one on the Arp3 subunit and one spanning Arp2 and ARPC1-but the importance of each site in the regulation of force-producing actin networks is unclear. Here, we identify mutations in budding yeast Arp2/3 complex that decrease or block engagement of Las17, the budding yeast WASP, at each site. As in the mammalian system, both sites are required for maximal activation in vitro. Dimerization of Las17 partially restores activity of mutations at both CA-binding sites. Arp2/3 complexes defective at either site assemble force-producing actin networks in a bead motility assay, but their reduced activity hinders motility by decreasing actin assembly near the bead surface and by failing to suppress actin filament bundling within the networks. While even the most defective Las17-binding site mutants assembled actin filaments at endocytic sites, they showed significant internalization defects, potentially because they lack the proper architecture to drive plasma membrane remodeling. Together, our data indicate that both Las17-binding sites are important to assemble functional endocytic actin networks in budding yeast, but Arp2/3 complex retains some activity in vitro and in vivo even with a severe defect at either Las17-binding site.

Controlled WASp activity regulates the proliferative response for Treg cell differentiation in the thymus.

The Wiskott-Aldrich syndrome protein (WASp) regulates actin cytoskeletal dynamics and function of hematopoietic cells. Mutations in the WAS gene lead to two different syndromes; Wiskott-Aldrich syndrome (WAS) caused by loss-of-function mutations, and X-linked neutropenia (XLN) caused by gain-of-function mutations. We previously showed that WASp-deficient mice have a decreased number of regulatory T (Treg) cells in the thymus and the periphery. We here evaluated the impact of WASp mutations on Treg cells in the thymus of WAS and XLN mouse models. Using in vitro Treg differentiation assays, WAS CD4 single-positive thymocytes have decreased differentiation to Treg cells, despite normal early signaling upon IL-2 and TGF-ß stimulation. They failed to proliferate and express CD25 at high levels, leading to poor survival and a lower number of Foxp3+ Treg cells. Conversely, XLN CD4 single-positive thymocytes efficiently differentiate into Foxp3+ Treg cells following a high proliferative response to IL-2 and TGF-ß, associated with high CD25 expression when compared with WT cells. Altogether, these results show that specific mutations of WASp affect Treg cell development differently, demonstrating a critical role of WASp activity in supporting Treg cell development and expansion.

LncRNA MYLK antisense RNA 1 activates cell division cycle 42/Neutal Wiskott-Aldrich syndrome protein pathway via microRNA-101-5p to accelerate epithelial-to-mesenchymal transition of colon cancer cells.

Long noncoding RNA MYLK antisense RNA 1 (MYLK-AS1) is the crux in multiple diseases. Therefore, the purpose of this study was to investigate the possible mechanism of MYLK-AS1. A total of 62 colon cancer (CC) specimens and paired adjacent normal tissues were collected, and the expression of MYLK-AS1, microRNA (miR)-101-5p/cell division cycle 42 (CDC42) was detected. CC cell lines were transfected with MYLK-AS1, miR-101-5p, CDC42-related plasmids, and the biological functions and markers of epithelial-mesenchymal transition (EMT) were analyzed. The binding relationship between MYLK-AS1, miR-101-5p, and CDC42 was evaluated. In CC tissues and cell lines, MYLK-AS1 and CDC42 were highly expressed, and miR-101-5p was lowly expressed. Inhibition of MYLK-AS1 or upregulation of miR-101-5p can inhibit CC cell growth and EMT. miR-101-5p inhibited CDC42/N-wasp axis activation in CC cells by targeting CDC42. Knockdown of CDC42 or upregulation of miR-101-5p partially reversed the effects caused by upregulation of MYLK-AS1. MYLK-AS1, which is significantly upregulated in CC, may be a molecular sponge for miR-101-5p, and MYLK-AS1 promotes the activation of the CDC42/N-wasp axis in CC cells by targeting CDC42 through miR-101-5p, which in turn promotes tumor development. MYLK-AS1 may be a potential biomarker and target for CC therapy.

Role of Wiskott Aldrich syndrome protein in haematological malignancies: genetics, molecular mechanisms and therapeutic strategies.

As patients continue to suffer from lymphoproliferative and myeloproliferative diseases known as haematopoietic malignancies can affect the bone marrow, blood, lymph nodes, and lymphatic and non-lymphatic organs. Despite advances in the current treatment, there is still a significant challenge for physicians to improve the therapy of HMs. WASp is an important regulator of actin polymerization and the involvement of WASp in transcription is thought to be linked to the DNA damage response and repair. In some studies, severe immunodeficiency and lymphoid malignancy are caused by WASp mutations or the absence of WASp and these mutations in WAS can alter the function and/or expression of the intracellular protein. Loss-of-function and Gain-of-function mutations in WASp have an impact on cancer malignancies' incidence and onset. Recent studies suggest that depending on the clinical or experimental situation, WASPs and WAVEs can operate as a suppressor or enhancers for cancer malignancy. These dual functions of WASPs and WAVEs in cancer likely arose from their multifaceted role in cells that could be targeted for anticancer drug development. The significant role and their association of WASp in Chronic myeloid leukaemia, Juvenile myelomonocytic leukaemia and T-cell lymphoma is discussed. In this review, we described the structure and function of WASp and its family mechanism, analysing major regulatory effectors and summarising the clinical relevance and drugs that specifically target WASp in disease treatment in various hematopoietic malignancies by different approaches.

Association of Wiskott-Aldrich syndrome protein (WASp) in epigenetic regulation of B cell differentiation in non-small-cell lung cancer (NSCLC).

Non-small-cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancer which is the deadliest type of cancer for both men and women. Previous studies already showed that cell-intrinsic loss of WASp causes B cell tolerance and WASp deficiency in T helper (TH) cells is linked to negative effects on cytokine gene transcription necessary for TH1 differentiation. In the current study, we investigated the molecular mechanisms involved in WASp-mediated epigenetic regulation of B cell differentiation during NSCLC. Our ChIP-qPCR data suggest the less percentage enrichment of the B cell differentiating factors (Ikaros, Pax5, PU.1, BATF) and WASp across the WAS gene in the B cells of NSCLC patients in comparison with normal healthy donors and overexpression of WASp showed the reverse effects. WASp-depleted B cells while co-culturing with respective PBMCs isolated from normal healthy donors and NSCLC patients, we observed upregulation of TH2-, TH17-, and Treg-specific cytokines (IL4, ILI7A, IL10) & transcription factors (GATA3, RORC, FOXP3) and downregulation of TH1-specific cytokine (IFNγ) & transcription factor (TBX21). Our study showed that the overexpression of WASp resulted into upregulation of B cell differentiating factors, tumor suppressor protein (p53), histone methylation marker (H3K4me3) with concomitant downregulation of tumor-promoting factors (Notch 1, ß-Catenin, DNAPKcs) and histone deacetylation marker (HDAC2) and increase in percentage cytotoxicity of NSCLC-specific cells (A549). Successful overexpression of WASp not only helps in epigenetic regulation of B cell differentiation but also supports tumor suppression in NSCLC. Thus, WASp can be targeted for therapeutic intervention of NSCLC.

Mechanism of synergistic activation of Arp2/3 complex by cortactin and WASP-family proteins.

Cortactin coactivates Arp2/3 complex synergistically with WASP-family nucleation-promoting factors (NPFs) and stabilizes branched networks by linking Arp2/3 complex to F-actin. It is poorly understood how cortactin performs these functions. We describe the 2.89 Å resolution cryo-EM structure of cortactin's N-terminal domain (Cort1-76) bound to Arp2/3 complex. Cortactin binds Arp2/3 complex through an inverted Acidic domain (D20-V29), which targets the same site on Arp3 as the Acidic domain of NPFs but with opposite polarity. Sequences N- and C-terminal to cortactin's Acidic domain do not increase its affinity for Arp2/3 complex but contribute toward coactivation with NPFs. Coactivation further increases with NPF dimerization and for longer cortactin constructs with stronger binding to F-actin. The results suggest that cortactin contributes to Arp2/3 complex coactivation with NPFs in two ways, by helping recruit the complex to F-actin and by stabilizing the short-pitch (active) conformation, which are both byproducts of cortactin's core function in branch stabilization.

Outcomes of hematopoietic stem cell gene therapy for Wiskott-Aldrich syndrome.

Wiskott-Aldrich syndrome (WAS) is a rare X-linked disorder characterized by combined immunodeficiency, eczema, microthrombocytopenia, autoimmunity, and lymphoid malignancies. Gene therapy (GT) to modify autologous CD34+ cells is an emerging alternative treatment with advantages over standard allogeneic hematopoietic stem cell transplantation for patients who lack well-matched donors, avoiding graft-versus-host-disease. We report the outcomes of a phase 1/2 clinical trial in which 5 patients with severe WAS underwent GT using a self-inactivating lentiviral vector expressing the human WAS complementary DNA under the control of a 1.6-kB fragment of the autologous promoter after busulfan and fludarabine conditioning. All patients were alive and well with sustained multilineage vector gene marking (median follow-up: 7.6 years). Clinical improvement of eczema, infections, and bleeding diathesis was universal. Immune function was consistently improved despite subphysiologic levels of transgenic WAS protein expression. Improvements in platelet count and cytoskeletal function in myeloid cells were most prominent in patients with high vector copy number in the transduced product. Two patients with a history of autoimmunity had flares of autoimmunity after GT, despite similar percentages of WAS protein-expressing cells and gene marking to those without autoimmunity. Patients with flares of autoimmunity demonstrated poor numerical recovery of T cells and regulatory T cells (Tregs), interleukin-10-producing regulatory B cells (Bregs), and transitional B cells. Thus, recovery of the Breg compartment, along with Tregs appears to be protective against development of autoimmunity after GT. These results indicate that clinical and laboratory manifestations of WAS are improved with GT with an acceptable safety profile. This trial is registered at clinicaltrials.gov as #NCT01410825.

The Wiskott-Aldrich syndrome protein is required for positive selection during T-cell lineage differentiation.

The Wiskott-Aldrich syndrome (WAS) is an X-linked primary immune deficiency caused by a mutation in the WAS gene. This leads to altered or absent WAS protein (WASp) expression and function resulting in thrombocytopenia, eczema, recurrent infections, and autoimmunity. In T cells, WASp is required for immune synapse formation. Patients with WAS show reduced numbers of peripheral blood T lymphocytes and an altered T-cell receptor repertoire. In vitro, their peripheral T cells show decreased proliferation and cytokine production upon aCD3/aCD28 stimulation. It is unclear whether these T-cell defects are acquired during peripheral activation or are, in part, generated during thymic development. Here, we assessed the role of WASp during T-cell differentiation using artificial thymic organoid cultures and in the thymus of humanized mice. Although CRISPR/Cas9 WAS knockout hematopoietic stem and progenitor cells (HSPCs) rearranged the T-cell receptor and differentiated to T-cell receptor (TCR)+ CD4+ CD8+ double-positive (DP) cells similar to wild-type HSPCs, a partial defect in the generation of CD8 single-positive (SP) cells was observed, suggesting that WASp is involved in their positive selection. TCR repertoire analysis of the DP and CD8+ SP population, however, showed a polyclonal repertoire with no bias toward autoreactivity. To our knowledge, this is the first study of the role of WASp in human T-cell differentiation and on TCR repertoire generation.

Wiskott-Aldrich syndrome protein interacts and inhibits diacylglycerol kinase alpha promoting IL-2 induction.

Background: Phosphorylation of diacylglycerol by diacylglycerol-kinases represents a major inhibitory event constraining T cell activation upon antigen engagement. Efficient TCR signalling requires the inhibition of the alpha isoform of diacylglycerol kinase, DGKα, by an unidentified signalling pathway triggered by the protein adaptor SAP. We previously demonstrated that, in SAP absence, excessive DGKα activity makes the T cells resistant to restimulation-induced cell death (RICD), an apoptotic program counteracting excessive T cell clonal expansion. Results: Herein, we report that the Wiskott-Aldrich syndrome protein (WASp) inhibits DGKα through a specific interaction of the DGKα recoverin homology domain with the WH1 domain of WASp. Indeed, WASp is necessary and sufficient for DGKα inhibition, and this WASp function is independent of ARP2/3 activity. The adaptor protein NCK-1 and the small G protein CDC42 connect WASp-mediated DGKα inhibition to SAP and the TCR signalosome. In primary human T cells, this new signalling pathway is necessary for a full response in terms of IL-2 production, while minimally affecting TCR signalling and restimulation-induced cell death. Conversely, in T cells made resistant to RICD by SAP silencing, the enhanced DAG signalling due to DGKα inhibition is sufficient to restore apoptosis sensitivity. Conclusion: We discover a novel signalling pathway where, upon strong TCR activation, the complex between WASp and DGKα blocks DGKα activity, allowing a full cytokine response.

Small GTPase Cdc42, WASP, and scaffold proteins for higher-order assembly of the F-BAR domain protein.

The higher-order assembly of Bin-amphiphysin-Rvs (BAR) domain proteins, including the FCH-BAR (F-BAR) domain proteins, into lattice on the membrane is essential for the formation of subcellular structures. However, the regulation of their ordered assembly has not been elucidated. Here, we show that the higher ordered assembly of growth-arrested specific 7 (GAS7), an F-BAR domain protein, is regulated by the multivalent scaffold proteins of Wiskott-Aldrich syndrome protein (WASP)/neural WASP, that commonly binds to the BAR domain superfamily proteins, together with WISH, Nck, the activated small guanosine triphosphatase Cdc42, and a membrane-anchored phagocytic receptor. The assembly kinetics by fluorescence resonance energy transfer monitoring indicated that the GAS7 assembly on liposomes started within seconds and was further increased by the presence of these proteins. The regulated GAS7 assembly was abolished by Wiskott-Aldrich syndrome mutations both in vitro and in cellular phagocytosis. Therefore, Cdc42 and the scaffold proteins that commonly bind to the BAR domain superfamily proteins promoted GAS7 assembly.

[Wiskott-Aldrich syndrome with platelets of normal size and c.295C>T mutation of the WAS gene. Case report]. / Síndrome de Wiskott-Aldrich con plaquetas de tamaño normal y mutación c.295C>T en el gen WAS. Informe de caso.

BACKGROUND: Wiskott-Aldrich syndrome is an Inborn Error of Immunity characterized by thrombocytopenia, small platelets, severe eczema, recurrent infections, tendency to autoimmune diseases and neoplasms. The diagnosis of the syndrome can be difficult, especially when platelets are of normal size. CASE REPORT: A three-year-old male patient was referred to a specialized sector of university hospital for presenting acute otitis media that progressed to sepsis by Haemophilus influenzae. At one month of age, he had been diagnosed with autoimmune thrombocytopenia, and splenectomy was performed at two years of age. During follow-up, three hospitalizations were necessary: an infection by Streptococcus pneumoniae, which progressed to sepsis; one due to exacerbation of eczema, isolating S. epidermidis; another due to fever of undetermined origin. The tests showed normal number of platelets after splenectomy, platelets always with normal size. At age four, tests were performed: IgE 3128 Ku/L; IgA, IgG, and normal anti-polysaccharide antibodies; decreased IgM; decrease CD19, TCD4, naïve T and B; increased TCD8; normal NK. A diagnostic hypothesis of "probable" WAS was made. Genetic research has identified the c.295C>T mutation in the WAS gene. CONCLUSIONS: The case reported expressed a new mutation in the SWA gene, characterized by clinical manifestations of the mild phenotype of Wiskott-Aldrich syndrome, with thrombocytopenia, platelets of normal size, and X-linked inheritance. It is important to establish the early diagnosis and treatment to offer a better quality of life in these patients.

ANTECEDENTES: El síndrome de Wiskott-Aldrich es un error innato de la inmunidad, distinguido por trombocitopenia, plaquetas pequeñas, eccema severo, infecciones recurrentes, y susceptibilidad a enfermedades autoinmunes y neoplasias. El diagnóstico es difícil de establecer, especialmente cuando las plaquetas son de tamaño normal. REPORTE DE CASO: Paciente masculino de 3 años, enviado al Hospital Universitario da Santa Casa de São Paulo, Brasil, por otitis media aguda, con evolución a sepsis por Haemophilus influenzae. Al mes de edad fue diagnosticado con trombocitopenia autoinmune, y a los 2 años se llevó a cabo explenectomía. Durante el seguimiento requirió tres hospitalizaciones: una por infección por Streptococcus pneumoniae, que evolucionó a sepsis; otra por exacerbación de eccema, aislándose S. epidermidis, y la última por fiebre de origen indeterminado. Las pruebas de laboratorio informaron: concentración de plaquetas dentro de los valores de referencia después de la esplenectomía, y de tamaño normal. A los 4 años se efectuaron nuevas pruebas, que reportaron: IgE 3128 kU/L; IgA, IgG y anticuerpos anti-polisacáridos normales; disminución de IgM y de CD19, TCD4, T y B vírgenes; aumento de TCD8; NK normales. Se sospechó el diagnóstico de síndrome de Wiskott-Aldrich. Mediante estudios de genética se identificó la mutación c.295C>T en el gen WAS. CONCLUSIONES: El caso aquí expuesto expresó una nueva mutación en el gen SWA, caracterizado por manifestaciones clínicas de fenotipo leve del síndrome de Wiskott-Aldrich, con trombocitopenia, plaquetas de tamaño normal y herencia ligada al cromosoma X. Es importante establecer el diagnóstico y tratamiento oportunos para ofrecer una mejor calidad de vida en estos pacientes.

GxcM-Fbp17/RacC-WASP signaling regulates polarized cortex assembly in migrating cells via Arp2/3.

The actin-rich cortex plays a fundamental role in many cellular processes. Its architecture and molecular composition vary across cell types and physiological states. The full complement of actin assembly factors driving cortex formation and how their activities are spatiotemporally regulated remain to be fully elucidated. Using Dictyostelium as a model for polarized and rapidly migrating cells, we show that GxcM, a RhoGEF localized specifically in the rear of migrating cells, functions together with F-BAR protein Fbp17, a small GTPase RacC, and the actin nucleation-promoting factor WASP to coordinately promote Arp2/3 complex-mediated cortical actin assembly. Overactivation of this signaling cascade leads to excessive actin polymerization in the rear cortex, whereas its disruption causes defects in cortical integrity and function. Therefore, apart from its well-defined role in the formation of the protrusions at the cell front, the Arp2/3 complex-based actin carries out a previously unappreciated function in building the rear cortical subcompartment in rapidly migrating cells.

A Novel Mutation Leading to Wiskott-Aldrich Syndrome in an Ethiopian Boy: a Case Report and a Review of Literature.

Wiskott-Aldrich syndrome is an X-linked recessive primary immune-deficiency disorder very rarely reported from black African children. A 12-year old boy with recurrent sinopulmonary and diarrheal infections, eczema, thrombocytopenia, and low platelet volume was found by whole genome sequencing to harbor a predicted pathogenic c.1205dupC (p.Pro403Alafs*92) variant of a mutation in the WAS gene - confirming the diagnosis. This case report summarizes his presentation and management and provides a useful summary of the diagnosis and the responsible novel genetic mutation.

N-WASP is competent for downstream signaling before full release from autoinhibition.

Allosteric regulation of intrinsically disordered proteins (IDPs) is still vastly understudied compared to the counterpart of structured proteins. Here, we used molecular dynamics simulations to characterize the regulation of the IDP N-WASP by the binding of its basic region with inter- and intramolecular ligands (PIP2 and an acidic motif, respectively). The intramolecular interactions keep N-WASP in an autoinhibited state; PIP2 binding frees the acidic motif for interacting with Arp2/3 and thereby initiating actin polymerization. We show that PIP2 and the acidic motif compete in binding with the basic region. However, even when PIP2 is present at 30% in the membrane, the acidic motif is free of contact with the basic region ("open" state) in only 8.5% of the population. The very C-terminal three residues of the A motif are crucial for Arp2/3 binding; conformations where only the A tail is free are present at a much higher population than the open state (40- to 6-fold, depending on the PIP2 level). Thus, N-WASP is competent for Arp2/3 binding before it is fully freed from autoinhibition.

Why Does Synergistic Activation of WASP, but Not N-WASP, by Cdc42 and PIP2 Require Cdc42 Prenylation?

Human WASP and N-WASP are homologous proteins that require the binding of multiple regulators, including the acidic lipid PIP2 and the small GTPase Cdc42, to relieve autoinhibition before they can stimulate the initiation of actin polymerization. Autoinhibition involves intramolecular binding of the C-terminal acidic and central motifs to an upstream basic region and GTPase binding domain. Little is known about how a single intrinsically disordered protein, WASP or N-WASP, binds multiple regulators to achieve full activation. Here we used molecular dynamics simulations to characterize the binding of WASP and N-WASP with PIP2 and Cdc42. In the absence of Cdc42, both WASP and N-WASP strongly associate with PIP2-containing membranes, through their basic region and also possibly through a tail portion of the N-terminal WH1 domain. The basic region also participates in Cdc42 binding, especially for WASP; consequently Cdc42 binding significantly compromises the ability of the basic region in WASP, but not N-WASP, to bind PIP2. PIP2 binding to the WASP basic region is restored only when Cdc42 is prenylated at the C-terminus and tethered to the membrane. This distinction in the activation of WASP and N-WASP may contribute to their different functional roles.

Branching out in different directions: Emerging cellular functions for the Arp2/3 complex and WASP-family actin nucleation factors.

The actin cytoskeleton impacts practically every function of a eukaryotic cell. Historically, the best-characterized cytoskeletal activities are in cell morphogenesis, motility, and division. The structural and dynamic properties of the actin cytoskeleton are also crucial for establishing, maintaining, and changing the organization of membrane-bound organelles and other intracellular structures. Such activities are important in nearly all animal cells and tissues, although distinct anatomical regions and physiological systems rely on different regulatory factors. Recent work indicates that the Arp2/3 complex, a broadly expressed actin nucleator, drives actin assembly during several intracellular stress response pathways. These newly described Arp2/3-mediated cytoskeletal rearrangements are coordinated by members of the Wiskott-Aldrich Syndrome Protein (WASP) family of actin nucleation-promoting factors. Thus, the Arp2/3 complex and WASP-family proteins are emerging as crucial players in cytoplasmic and nuclear activities including autophagy, apoptosis, chromatin dynamics, and DNA repair. Characterizations of the functions of the actin assembly machinery in such stress response mechanisms are advancing our understanding of both normal and pathogenic processes, and hold great promise for providing insights into organismal development and interventions for disease.

The proline-rich domain of fission yeast WASp (Wsp1p) interacts with actin filaments and inhibits actin polymerization.

Members of the Wiskott-Aldrich Syndrome protein (WASp) family activate Arp2/3 complex (actin-related proteins 2 and 3 complex) to form actin filament branches. The proline-rich domain (PRD) of WASp contributes to branching nucleation, and the PRD of budding yeast Las17 binds actin filaments [Urbanek AN et al. (2013) Curr Biol 23, 196-203]. Biochemical assays showed the recombinant PRD of fission yeast Schizosaccharomyces pombe Wsp1p binds actin filaments with micromolar affinity. Recombinant PRDs of both Wsp1p and Las17p slowed the elongation of actin filaments by Mg-ATP-actin monomers by half and slowed the spontaneous polymerization of Mg-ATP-actin monomers modestly. The affinity of PRDs of WASp-family proteins for actin filaments is high enough to contribute to the reported stimulation of actin filament branching by Arp2/3 complex.