Brad A. Bryan

What tangled webs they weave: Rho-GTPase control of angiogenesis

Abstract.The members of the Rho family of small GTPases are involved in an array of cellular processes, including regulation of the actin cytoskeleton, cell polarity, microtubule dynamics, membrane transport, and transcription factor activity. Recent findings have implicated the Rho-proteins as key regulators of angiogenesis, modulating a diversity of cellular processes, including vascular permeability, extracellular matrix remodeling, migration, proliferation, morphogenesis, and survival.

The function of vascular endothelial growth factor

Vascular endothelial growth factor (VEGF) is considered the master regulator of angiogenesis during growth and development, as well as in disease states such as cancer, diabetes, and macular degeneration. This review details our current understanding of VEGF signaling and discusses the benefits and unexpected side effects of promising anti‐angiogenic therapeutics that are currently being used to inhibit neovacularization in tumors.

Anti-angiogenic therapy: adapting strategies to overcome resistant tumors.

Healthy cells, as well as benign and malignant tumors, depend upon the bodys blood supply to bring in oxygen and nutrients and carry away waste products. Using this property against tumors, anti‐angiogenic therapy targets the tumor vasculature with the aim of starving the tumor, and has demonstrated exceptional clinical efficacy against a number of tumors. This review discusses the current state of knowledge regarding anti‐angiogenic therapies presently available to patients, and garners from both preclinical and clinical literature the benefits and side effects associated with anti‐angiogenic therapies, the unfortunate mechanisms of acquired resistance to these novel therapeutics, and highlights promising next generation anti‐angiogenics that may overcome the limitations encountered with first generation therapies. J. Cell. Biochem. 111: 543–553, 2010.

The Rho-family guanine nucleotide exchange factor GEFT enhances retinoic acid- and cAMP-induced neurite outgrowth

The Rho GTPases are important regulators of neurite outgrowth and pathfinding. We have recently reported that a Rho‐family guanine nucleotide exchange factor, GEFT, modulates dendrite spine morphology and basal neurite outgrowth in primary hippocampal neurons and Neuro2A cells, respectively. Here we demonstrate that GEFT protein is highly expressed in all regions of the brain and is highly up‐regulated upon treatment of Neuro2A cells with retinoic acid and dibutyric cAMP, which promote dendrite and axon‐like neurite extensions, respectively. Within retinoic acid‐induced neurite extensions, GEFT is localized to actin‐enriched regions in the primary neurites, with little or no expression from secondary branches. Dibutyric cAMP‐induced neurite extensions are highly concentrated for GEFT at the actin‐rich distal tip of the growth cone. Additionally, we demonstrate that GEFT promotes neurite outgrowth in undifferentiated as well as differentiated Neuro2A cells. Together, our data provide new evidence suggesting that GEFT is an important regulator of multiple processes involved in axon and dendrite formation.

Rock1 & 2 Perform Overlapping and Unique Roles in Angiogenesis and Angiosarcoma Tumor Progression

The serine/threonine protein kinase paralogs ROCK1 & 2 have been implicated as essential modulators of angiogenesis; however their paralog-specific roles in endothelial function are unknown. shRNA knockdown of ROCK1 or 2 in endothelial cells resulted in a significant disruption of in vitro capillary network formation, cell polarization, and cell migration compared to cells harboring non-targeting control shRNA plasmids. Knockdowns led to alterations in cytoskeletal dynamics due to ROCK1 & 2-mediated reductions in actin isoform expression, and ROCK2-specific reduction in myosin phosphatase and cofilin phosphorylation. Knockdowns enhanced cell survival and led to ROCK1 & 2-mediated reduction in caspase 6 and 9 cleavage, and a ROCK2-specific reduction in caspase 3 cleavage. Microarray analysis of ROCK knockdown lines revealed overlapping and unique control of global transcription by the paralogs, and a reduction in the transcriptional regulation of just under 50% of VEGF responsive genes. Finally, paralog knockdown in xenograft angiosarcoma tumors resulted in a significant reduction in tumor formation. Our data reveals that ROCK1 & 2 exhibit overlapping and unique roles in normal and dysfunctional endothelial cells, that alterations in cytoskeletal dynamics are capable of overriding mitogen activated transcription, and that therapeutic targeting of ROCK signaling may have profound impacts for targeting angiogenesis.

Nutrient-specific effects in the coordination of cell growth with cell division in continuous cultures of Saccharomyces cerevisiae

Cell cycle progression of Saccharomyces cerevisiae cells was monitored in continuous cultures limited for glucose or nitrogen. The G1 cell cycle phase, before initiation of DNA replication, did not exclusively expand when growth rate decreased. Especially during nitrogen limitation, non-G1 phases expanded almost as much as G1. In addition, cell size remained constant as a function of growth rate. These results contrast with current views that growth requirements are met before initiation of DNA replication, and suggest that distinct nutrient limitations differentially impinge on cell cycle progression.

The UV Response in Saccharomyces cerevisiae Involves the Mitogen-Activated Protein Kinase Slt2p

Exposure to UV causes a response in yeast and mammalian cells, which is distinct from the response to DNA damage. We report that the mitogen-activated protein kinase Slt2p is involved in this response in Saccharomyces cerevisiae. Thus, budding yeast and mammalian cells respond to UV by using very similar signal transduction pathways.

Aberrations in Angiogenic Signaling and MYC Amplifications are Distinguishing Features of Angiosarcoma.

Angiosarcomas are very aggressive, rare malignant tumors that originate from vascular or lymphatic vessels and primarily occur following chemical exposure or radiation therapy. Tumor response to either chemotherapy, radiation, or novel anti-angiogenic therapeutics is very low, and because little is known regarding the aberrant signaling that controls these tumors, personalized treatment options for many of these patients are lacking. In this review, we summarize several recent findings regarding the genomics of angiosarcomas, including new discoveries regarding aberrant angiogenic signaling and Myc amplification as key features of this tumor type.

Evidence for control of nitrogen metabolism by a START-dependent mechanism in Saccharomyces cerevisiae

It is generally thought that cell growth and metabolism regulate cell division and not vice versa. Here, we examined Saccharomyces cerevisiae cells growing under conditions of continuous culture in a chemostat. We found that loss of G1 cyclins, or inactivation of the cyclin-dependent kinase Cdc28p, reduced the activity of glutamate synthase (Glt1p), a key enzyme in nitrogen assimilation. We also present evidence indicating that the G1 cyclin-dependent control of Glt1p may involve Jem1p, a DnaJ-type chaperone. Our results suggest that completion of START may be linked to nitrogen metabolism.

Everolimus affects vasculogenic mimicry in renal carcinoma resistant to sunitinib

Angiogenesis is hallmark of clear cell renal cell carcinogenesis. Anti-angiogenic therapies have been successful in improving disease outcome; however, most patients treated with anti-angiogenic agents will eventually progress. In this study we report that clear cell renal cell carcinoma was associated with vasculogenic mimicry in both mice and human with tumor cells expressing endothelial markers in the vicinity of tumor vessels. We show that vasculogenic mimicry was efficiently targeted by sunitinib but eventually associated with tumor resistance and a more aggressive phenotype both in vitro and in vivo. Re-challenging these resistant tumors in mice, we showed that second-line treatment with everolimus particularly affected vasculogenic mimicry and tumor cell differentiation compared to sorafenib and axitinib. Finally, our results highlighted the phenotypic and genotypic changes at the tumor cell and microenvironment levels during sunitinib response and progression and the subsequent improvement second-line therapies bring to the current renal cell carcinoma treatment paradigm.