The yeast retrovirus-like element Ty3 GAG3 gene encodes a Gag3 polyprotein analogous to retroviral Gag. Gag3 lacks matrix, but contains capsid, spacer, and nucleocapsid domains. Expression of a Ty3 Gag3 or capsid domain optimized for expression in Escherichia coli was sufficient for Ty3 particle assembly. Virus-like ordered particles assembled from Gag3 were similar in size to immature particles from yeast and contained nucleic acid. However, particles assembled from the CA domain were variable in size and displayed much less organization than native particles. These results indicate that assembly can be driven through interactions among capsid subunits in the particle, but that the nucleocapsid domain, likely in association with RNA, confers order upon this process.

Cancer cells derived from Glioblastoma multiforme possess membranous protrusions allowing these cells to infiltrate surrounding tissue, while resisting lymphocyte cytotoxicity. Microvilli and filopodia are supported by actin filaments cross-linked by fascin. Fascin-1 was genetically silenced within human U251 glioma cells; these knock-down glioma cells lost their microvilli/filopodia. The doubling time of these fascin-1 knock-down cells was doubled that of shRNA control U251 cells. Fascin-1 knock-down cells lost their transmigratory ability responding to interleukin-6 or insulin-like growth factor-1. Fascin-1 silenced U251 cells were more easily killed by cytolytic lymphocytes. Fascin-1 knock-down provides unique opportunities to augment glioma immunotherapy by simultaneously targeting several key glioma functions: like cell transmigration, cell division and resisting immune responses.

Gliomas are invasive cancers that resist all forms of attempted therapy. Immunotherapy using Ag-pulsed dendritic cells has improved survival in some patients. We present evidence that another level of complexity may also contribute to lack of responses by the lymphocytes toward gliomas. Atomic force microscopy of four different glioma types-human U251 and rat T9 and F98 glioma cells, including freshly isolated human glioblastoma multiforme neurosphere cultures (containing "stem cell-like cells")-revealed a complex surface topography with numerous microvilli and filopodia. These structures were not found on other cell types. Electron microscopy and immunofluorescence microscopy of glioma cells confirmed that microvilli are present. U251 cells with microvilli resisted the cytolytic actions of different human effector cells, (lymphokine-activated killer cells, γδ T cells, conventional CTLs, and chimeric Ag-receptor-redirected T cells) better than their nonmicrovilli-expressing counterparts. Killer lymphocytes released perforin, which was detected within the glioma's microvilli/filopodia, indicating these structures can receive the cytolytic effector molecules, but cytotoxicity is suboptimal. Air-dried gliomas revealed nodes within the microvilli/filopodia. The microvilli that penetrated 0.4-μm transwell chamber's pores resisted the actions of CTLs and physical damage. Those nodelike structures may represent a compartmentalization that resists physical damage. These microvilli may play multiple roles in glioma biology, such as invasion and resistance to lymphocyte-mediated killing.