Cancer stem cells (CSCs), a small and elusive population of undifferentiated cancer cells within tumors that drive tumor growth and recurrence, are believed to resemble normal stem cells. Although surrogate markers have been identified and compelling CSC theoretical models abound, actual proof for the existence of CSCs can only be had retrospectively. Hence, great store has come to be placed in isolating CSCs from cancers for in-depth analysis. On the other hand, although induced pluripotent stem cells (iPSCs) hold great promise for regenerative medicine, concern exists over the inadvertent co-transplantation of partially or undifferentiated stem cells with tumorigenic capacity. Here we demonstrate that the introduction of defined reprogramming factors (OCT4, SOX2, Klf4 and c-Myc) into MCF-10A nontumorigenic mammary epithelial cells, followed by partial differentiation, transforms the bulk of cells into tumorigenic CD44+/CD24low cells with CSC properties, termed here as induced CSC-like-10A or iCSCL-10A cells. These reprogrammed cells display a malignant phenotype in culture and form tumors of multiple lineages when injected into immunocompromised mice. Compared with other transformed cell lines, cultured iCSCL-10A cells exhibit increased resistance to the chemotherapeutic compounds, Taxol and Actinomycin D, but higher susceptibility to the CSC-selective agent Salinomycin and the Pin1 inhibitor Juglone. Restored expression of the cyclin-dependent kinase inhibitor p16INK4a abrogated the CSC properties of iCSCL-10A cells, by inducing cellular senescence. This study provides some insight into the potential oncogenicity that may arise via cellular reprogramming, and could represent a valuable in vitro model for studying the phenotypic traits of CSCs per se.

Parkinson disease (PD) is a relatively common neurodegenerative disorder that is characterized by the loss of dopaminergic neurons and by the formation of Lewy bodies (LBs), which are cytoplasmic inclusions containing aggregates of ␣-synuclein. Although certain post-translational modifications of ␣-synuclein and its related proteins are implicated in the genesis of LBs, the specific molecular mechanisms that both regulate these processes and initiate subsequent inclusion body formation are not yet well understood. We demonstrate in our current study, however, that the prolyl-isomerase Pin1 localizes to the LBs in PD brain tissue and thereby enhances the formation of ␣-synuclein immunoreactive inclusions. Immunohistochemical analysis of brain tissue from PD patients revealed that Pin1 localizes to 50 -60% of the LBs that show an intense halo pattern resembling that of ␣-synuclein. By utilizing a cellular model of ␣-synuclein aggregation, we also demonstrate that, whereas Pin1 overexpression facilitates the formation of ␣-synuclein inclusions, dominant-negative Pin1 expression significantly suppresses this process. Consistent with these observations, Pin1 overexpression enhances the protein half-life and insolubility of ␣-synuclein. Finally, we show that Pin1 binds synphilin-1, an ␣-synuclein partner, via its Ser-211-Pro and Ser-215-Pro motifs, and enhances its interaction with ␣-synuclein, thus likely facilitating the formation of ␣-synuclein inclusions. These results indicate that Pin1-mediated prolyl-isomerization plays a pivotal role in a post-translational modification pathway for ␣-synuclein aggregation and in the resultant Lewy body formations in PD. Parkinson disease (PD)2 is one of the most common neurodegenerative disorders and is characterized by the loss of dopaminergic neurons in the substantia nigra and by the presence of cytoplasmic inclusions known as Lewy bodies (LBs) in surviving neurons (1, 2). LBs have classically been considered as a pathological hallmark of PD, consisting of many components, including ␣-synuclein, which is one of the major constituents (3, 4). The first indication of a pathogenic role for ␣-synuclein in PD came from the results of linkage analysis of mutations in its gene in autosomal dominant forms of the disease (5, 6). ␣-Synuclein is an unfolded protein in its native state, but in a pathological state it can be induced to form either ␣-helical or ␤-sheet structures that result in the formation of insoluble ␣-synuclein aggregates (7,8). The aggregation of ␣-synuclein can be modified by a range of factors, both in vitro and in vivo, including environmental regulators of pH, temperature, ionic strength, and oxidative stress and by intrinsic intracellular pathways (8). The latter of these regulatory networks includes several ␣-synuclein-binding proteins such as synphilin-1 (9) and posttranslational modifications of related molecules, such as phosphorylation and ubiquitination (10 -12). Synphilin-1 was identified as a protein that interacts with ␣-synuclein and has been shown t...

Human immunodeficiency virus type 1 (HIV-1) utilizes the macromolecular machinery of the infected host cell to produce progeny virus. The discovery of cellular factors that participate in HIV-1 replication pathways has provided further insight into the molecular basis of virus-host cell interactions. Here, we report that the suppressor of cytokine signaling 1 (SOCS1) is an inducible host factor during HIV-1 infection and regulates the late stages of the HIV-1 replication pathway. SOCS1 can directly bind to the matrix and nucleocapsid regions of the HIV-1 p55 Gag polyprotein and enhance its stability and trafficking, resulting in the efficient production of HIV-1 particles via an IFN signaling-independent mechanism. The depletion of SOCS1 by siRNA reduces both the targeted trafficking and assembly of HIV-1 Gag, resulting in its accumulation as perinuclear solid aggregates that are eventually subjected to lysosomal degradation. These results together indicate that SOCS1 is a crucial host factor that regulates the intracellular dynamism of HIV-1 Gag and could therefore be a potential new therapeutic target for AIDS and its related disorders.AIDS ͉ pathogenesis ͉ drug target ͉ lysozyme

This study indicated that H19 was associated with stem cell phenotype in ALDH1-positive breast cancer. H19 regulates CSC and is associated with poor prognosis in breast cancer patients, particularly in triple-negative subtype.

The ongoing COVID-19 pandemic is a major global public health concern. Although rapid point-of-care testing for detecting viral antigen is important for management of the outbreak, the current antigen tests are less sensitive than nucleic acid testing. In our current study, we produce monoclonal antibodies (mAb) that exclusively react with SARS-CoV-2 and exhibit no cross-reactivity with other human coronaviruses including SARS-CoV. Molecular modeling suggest that the mAbs bind to epitopes present on the exterior surface of the nucleocapsid, making them suitable for detecting SARS-CoV-2 in clinical samples. We further select the optimal pair of anti-SARS-CoV-2 NP mAbs using ELISA, and then use this mAb pair to develop immunochromatographic assay augmented with silver amplification technology. Our mAbs recognize the variants of concern (501Y.V1-V3) that are currently in circulation. Due to their high performance, the mAbs of this study can serve as good candidates for developing antigen detection kits for COVID-19.

The death-associated protein Daxx is a multifunctional factor that regulates a variety of cellular processes, including transcription and apoptosis. Several previous reports have indicated that Daxx is induced upon oxidative stress and is then subjected to phosphorylation-based functional modification. However, the precise molecular events underlying these phosphorylation events remain largely unknown. We report in our current study that the peptidyl-prolyl isomerase Pin1 is highly overexpressed in malignant human gliomas and inhibits Daxx-mediated cellular apoptosis. The targeted inhibition of Pin1 by small interfering RNA in A172 glioblastoma cells significantly enhances the apoptotic response induced by hydrogen peroxide or stimulatory Fas antibodies. This is in turn accompanied by the increased induction of Daxx and the activation of the apoptosis signal-regulating kinase 1/c-Jun N-terminal kinase pathway. Furthermore, Pin1 binds to the phosphorylated Ser 178 -Pro motif in the Daxx protein, and Pin1 overexpression results in the rapid degradation of Daxx via the ubiquitin-proteasome pathway. Moreover, a Daxx-S178A mutant, which cannot interact with Pin1, demonstrates higher proapoptotic activity and is refractory to Pin1-mediated antiapoptotic effects. We further found that the expression levels of Pin1 inversely correlate with the degree of Daxx nuclear accumulation in human glioblastoma tissues. These results together indicate that Pin1-mediated prolyl isomerization plays an important role in the negative regulation of Daxx and thereby inhibits the oxidative stress-induced cellular apoptotic response, particularly in malignant tumor cells where Pin1 is often overexpressed.

The prominent characteristics of pluripotent stem cells are their unique capacity to self-renew and pluripotency. Although pluripotent stem cell proliferation is maintained by specific intracellular phosphorylation signaling events, it has not been well characterized how the resulting phosphorylated proteins are subsequently regulated. We here report that the peptidylprolyl isomerase Pin1 is indispensable for the self-renewal and maintenance of pluripotent stem cells via the regulation of phosphorylated Oct4 and other substrates. Pin1 expression was found to be up-regulated upon the induction of induced pluripotent stem (iPS) cells, and the forced expression of Pin1 with defined reprogramming factors was observed to further enhance the frequency of iPS cell generation. The inhibition of Pin1 activity significantly suppressed colony formation and induced the aberrant differentiation of human iPS cells as well as murine ES cells. We further found that Pin1 interacts with the phosphorylated Ser 12 -Pro motif of Oct4 and that this in turn facilitates the stability and transcriptional activity functions of Oct4. Our current findings thus uncover an atypical role for Pin1 as a putative regulator of the induction and maintenance of pluripotency via the control of phosphorylation signaling. These data suggest that the manipulation of Pin1 function could be a potential strategy for the stable induction and proliferation of human iPS cells.Stem cells are characterized by their ability to self-renew through mitotic cell division and to differentiate into a diverse range of specialized cell types (1, 2). Human pluripotent stem cell proliferation is maintained through the action of several transcription factors including Oct4 (octamer 4), SOX2, Klf-4, Nanog, and c-Myc, which perform reprogramming functions under the stimulatory effects of stem cell-specific growth factors, including basic fibroblast growth factor (3-5). Basic fibroblast growth factor signaling has been shown to be essential for pluripotency as its depletion from cell culture media leads to aberrant cell differentiation and cell death (6, 7). Fibroblast growth factors produce mitogenic effects in targeted cells via signaling through cell surface receptor tyrosine kinases (8). These kinases can initiate intracellular signaling in cells, which is transmitted and diffused by tyrosine phosphorylation of the assembled proteins and of cellular substrates, including protein kinases with specificity for serine/threonine residues (8, 9). Although this intracellular phosphorylation signaling might indeed contribute to the self-renewal and pluripotency of stem cells (10, 11), it has not yet been fully determined how these phosphorylated proteins are further regulated.Protein phosphorylation is a fundamental mode of intracellular signal transduction in a variety of key cellular processes such as cell proliferation, differentiation, and morphogenesis (12). A pivotal signaling mechanism that controls the function of phosphorylated proteins is the cis-trans isomerization of...

Cancer stem cells (CSCs) retain the capacity to propagate themselves through self-renewal and to produce heterogeneous lineages of cancer cells constituting the tumor. Novel drugs that target CSCs can potentially eliminate the tumor initiating cell population therefore resulting in complete cure of the cancer. We recently established a CSC-like model using induced pluripotent stem cell (iPSC) technology to reprogram and partially differentiate human mammary epithelial MCF-10A cells. Using the induced CSC-like (iCSCL) model, we developed a phenotypic drug assay system to identify agents that inhibit the stemness and self-renewal properties of CSCs. The selectivity of the agents was assessed using three distinct assays characterized by cell viability, cellular stemness and tumor sphere formation. Using this approach, we found that withaferin A (WA), an Ayurvedic medicine constituent, was a potent inhibitor of CSC stemness leading to cellular senescence primarily via the induction of p21Cip1 expression. Moreover, WA exhibited strong anti-tumorigenic activity against the iCSCL. These results indicate that our iCSCL model provides an innovative high throughput platform for a simple, easy, and cost-effective method to search for novel CSC-targeting drugs. Furthermore, our current study identified WA as a putative drug candidate for abrogating the stemness and tumor initiating ability of CSCs.