Chmp 1A is a mediator of the anti-proliferative effects of All-trans Retinoic Acid in human pancreatic cancer cells

<p>Abstract</p> <p>Background</p> <p>We recently have shown that Charged multivesicular protein/Chromatin modifying protein1A (Chmp1A) functions as a tumor suppressor in human pancreatic tumor cells. Pancreatic cancer has the worst prognosis of all cancers with a dismal 5-year survival rate. Preclinical studies using ATRA for treating human pancreatic cancer suggest this compound might be useful for treatment of pancreatic cancer patients. However, the molecular mechanism by which ATRA inhibits growth of pancreatic cancer cells is not clear. The objective of our study was to investigate whether Chmp1A is involved in ATRA-mediated growth inhibition of human pancreatic tumor cells.</p> <p>Results</p> <p>We performed microarray studies using HEK 293T cells and discovered that Chmp1A positively regulated Cellular retinol-binding protein 1 (CRBP-1). CRBP-1 is a key regulator of All-trans retinoic acid (ATRA) through ATRA metabolism and nuclear localization. Since our microarray data indicates a potential involvement of Chmp1A in ATRA signaling, we tested this hypothesis by treating pancreatic tumor cells with ATRA <it>in vitro</it>. In the ATRA-responsive cell lines, ATRA significantly increased the protein expression of Chmp1A, CRBP-1, P53 and phospho-P53 at serine 15 and 37 position. We found that knockdown of Chmp1A via shRNA abolished the ATRA-mediated growth inhibition of PanC-1 cells. Also, Chmp1A silencing diminished the increase of Chmp1A, P53 and phospho-P53 protein expression induced by ATRA. In the ATRA non-responsive cells, ATRA did not have any effect on the protein level of Chmp1A and P53. Chmp1A over-expression, however, induced growth inhibition of ATRA non-responsive cells, which was accompanied by an increase of Chmp1A, P53 and phospho-P53. Interestingly, in ATRA responsive cells Chmp1A is localized to the nucleus, which became robust upon ATRA treatment. In the ATRA-non-responsive cells, Chmp1A was mainly translocated to the plasma membrane upon ATRA treatment.</p> <p>Conclusion</p> <p>Collectively our data provides evidence that Chmp1A mediates the growth inhibitory activity of ATRA in human pancreatic cancer cells via regulation of CRBP-1. Our results also suggest that nuclear localization of Chmp1A is important in mediating ATRA signaling.</p

The Wingless Signaling Pathway Is Directly Involved inDrosophilaHeart Development

AbstractHeart development in both vertebrates andDrosophilais initiated by bilaterally symmetrical primordia that may be of equivalent embryological origin: the anterior lateral plate mesoderm in vertebrates and the dorsal-most mesoderm in arthropods. These mesodermal progenitors then merge into a heart tube at the ventral midline (vertebrates) or the dorsal midline (Drosophila). These observations suggest that there may be similarities between vertebrate and invertebrate heart development. The homeobox gene,tinman,is required for heart as well as visceral mesoderm formation inDrosophila,and at least one of several vertebrate genes with similarities in protein sequence and cardiac expression totinmanis crucial for heart development in vertebrates. Inductive signals are also required forDrosophilaheart development: The secreted gene product ofwingless(wg) is critical for heart development during a time period distinct from its function in segmentation and neurogenesis. Here, we show thatwgis epistatic to hedgehog (hh), another secreted segmentation gene product, in its requirement for heart formation. We also provide evidence show that downstream ofwgin the signal transduction cascade,dishevelled(dsh,a pioneer protein) andarmadillo(arm, β-catenin homolog) are mediating the cardiogenic Wg signal. In particular, overexpression ofdshcan restore heart formation in the absence ofwgfunction. We discuss the possibility that Wg signaling is part of a combinatorial mechanism to specify the cardiac mesoderm

Inductive and lineage mechanisms during heart development of Drosophila.

Drosophila heart development is initiated by bilaterally symmetrical primordia to form a simple tubular structure with two rows of cells. tinman, a homeobox gene, is absolutely required for heart and visceral muscle formation in Drosophila. Several vertebrate tinman relatives are predominately expressed in the early cardiac and pharyngeal endoderm progenitors, suggesting that there may be similarities between vertebrate and invertebrate heart development. Using Drosophila as a model system, I have shown that vertebrate tinman-related genes can readily replace Drosophila tinman in promoting visceral mesoderm but not heart development. However, another Drosophila homeobox gene, bagpipe, which is needed only for visceral muscle development, can not replace tinman. These data indicate that the functional equivalence of the tinman genes is specific to this group. Cell-cell communications via inductive signals from ectodermal to mesodermal germ layer play roles in patterning mesoderm. By employing a genetic approach, I have shown that the secreted gene product of wingless as well as its responsive genes is critical for heart development during a distinct time period. A serine threonine kinase, zeste-white 3/shaggy (known to be a negative wg signal mediator in other developmental processes), may play a dual role in Drosophila mesoderm development: it appears to act both as a positive regulator of dorsal mesoderm formation and a negative component of the cardiogenic wg cascade. Additionally, I have shown that Notch does not inhibit wg-mediated cardiogenesis. Rather, constitutively activated Notch appears to repress mesoderm differentiation. Many neural precursors undergo fixed, asymmetric cell divisions, a process that involves the genetic interaction of the localized determinant encoded by numb with sanpodo, whose product may be cytoskeleton-associated, and with Notch, to control the distinction between daughter cell fates. Here, I have shown that similar interactions also specify cell fates in the mesoderm by determining the identities of muscle founders as well as of heart precursors. Thus, asymmetric cell divisions, in addition to inductive mechanisms and transcriptional regulation, appear to play a major role in distinguishing between alternative cell fates for mesodermal patterning.Ph.D.Biological SciencesMolecular biologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/131065/2/9825317.pd

Heart development in Drosophila and vertebrates: Conservation of molecular mechanisms

Vertebrate and insect ( Drosophila ) hearts look and function quite differently from each other. Nevertheless, during embryogenesis their mesodermal origin and initial assembly into a linear heart tube are comparable in many respects. In the past few years, numerous gene functions have been identified that are utilized by both vertebrates and Drosophila for the specification and differentiation of the heart progenitor cells. These studies have begun with the discovery of the homeobox gene tinman in Drosophila and its vertebrate counterparts. By now, there is also evidence that MEF2 transcription factors and TGF-β signaling have cardiogenic functions in both these systems. Perhaps in a few years, the GATA and HAND transcription factors and Wnt signaling, which currently only have a demonstrated cardiogenic function in one of the systems, may also be part of this group. One of the pressing but still wide open questions is if the spectrum of targets for these transcription factors and signaling pathways is also conserved. Dev. Genet. 22:181–186, 1998. © 1998 Wiley-Liss, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/34685/1/1_ftp.pd

Anacardic acid inhibits pancreatic cancer cell growth, and potentiates chemotherapeutic effect by Chmp1A - ATM - p53 signaling pathway

Abstract Background Pancreatic cancer is one of the leading causes of cancer related death and its incidence has risen steadily. Although anticancer drugs have been developed based on the new molecular findings, the drugs have produced unsatisfactory results due to toxicity and resistance. Thus, a complementary therapeutic intervention is urgently needed for pancreatic cancer patients. Methods The aim of this study was to assess the potential therapeutic effect of Anacardic acid on pancreatic cancer in vitro and elucidate its underlying mechanisms. Human pancreatic cancer cells were treated with Anacardic acid and assessed for the cytotoxic effect using MTT and spheroid formation assays. Using the same methods, the synergy between Anacardic acid and 5-Fluorouracil or Gemcitabine was determined. To elucidate the underlying molecular mechanisms, Western blot analysis and immunocytochemistry were performed on cancer cells treated with Anacardic acid alone or in combination with 5-Fluorouracil or Gemcitabine. Chromatin Modifying Protein 1A (Chmp1A), Ataxia Telangiectasia Mutated (ATM), and p53 were the primary signaling molecules examined. In addition, Chmp1A was silenced with shRNA to examine the necessity of Chmp1A for the anticancer effect of Anacardic acid, 5-Fluorouracil, or Gemcitabine. Results Anacardic acid induced an anticancer effect in pancreatic cancer cell lines in a dose dependent manner, and increased the cytotoxicity of 5-Fluorouracil or Gemcitabine in MTT cell viability assays. In spheroid formation assays, spheroids formed were smaller in size and in number upon Anacardic acid treatment compared to control. Mechanistically, Anacardic acid exerted its anticancer activity via the activation of Chmp1A, ATM, and p53. Interestingly, 5-Fluorouracil and Gemcitabine also induced an increase in Chmp1A protein level, suggesting that Chmp1A might mediate the cytotoxic action of chemotherapeutics. Silencing experiments indicate that Chmp1A is required for the action of Anacardic acid, but not for 5-Fluorouracil or Gemcitabine. Conclusions Our data suggests that Anacardic Acid might be a promising complementary supplement to slow the initiation or progression of pancreatic cancer