Absence of XMRV and Closely Related Viruses in Primary Prostate Cancer Tissues Used to Derive the XMRV-Infected Cell Line 22Rv1

The 22Rv1 cell line is widely used for prostate cancer research and other studies throughout the world. These cells were established from a human prostate tumor, CWR22, that was serially passaged in nude mice and selected for androgen independence. The 22Rv1 cells are known to produce high titers of xenotropic murine leukemia virus-related virus (XMRV). Recent studies suggested that XMRV was inadvertently created in the 1990's when two murine leukemia virus (MLV) genomes (pre-XMRV1 and pre-XMRV-2) recombined during passaging of the CWR22 tumor in mice. The conclusion that XMRV originated from mice and not the patient was based partly on the failure to detect XMRV in early CWR22 xenografts. While that deduction is certainly justified, we examined the possibility that a closely related virus could have been present in primary tumor tissue. Here we report that we have located the original prostate tumor tissue excised from patient CWR22 and have assayed the corresponding DNA by PCR and the tissue sections by fluorescence in situ hybridization for the presence of XMRV or a similar virus. The primary tumor tissues lacked mouse DNA as determined by PCR for intracisternal A type particle DNA, thus avoiding one of the limitations of studying xenografts. We show that neither XMRV nor a closely related virus was present in primary prostate tissue of patient CWR22. Our findings confirm and reinforce the conclusion that XMRV is a recombinant laboratory-generated mouse virus that is highly adapted for human prostate cancer cells

Retrotransposons and the evolution of mammalian gene expression

Transposable elements, and retroviral-like elements in particular, are a rich potential source of genetic variation within a host's genome. Many mutations of endogenous genes in phylogenetically diverse organisms are due to insertion of elements that affect gene expression by altering the normal pattern of regulation. While few such associations are known to have been maintained over time, two recently elucidated examples suggest transposable elements may have a significant impact in evolution of gene expression. The first example, concerning the mouse sex-limited protein ( Slp ), clearly establishes that ancient retroviral enhancer sequences now confer hormonal dependence on the adjacent gene. The second example shows that within the human amylase gene family, salivary specific expression has arisen due to inserted sequences, deriving perhaps from a conjunction of two retrotransposable elements.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/42800/1/10709_2004_Article_BF00133720.pd

Sequence and expression of a novel GABAA receptor alpha subunit

Cloned cDNA encoding the bovine alpha 4 subunit of the GABAA receptor has been isolated. The predicted 51 amino acid long mature protein contains an exceptionally long intracellular domain and shares 53-56% sequence similarity to the previously characterized alpha 1, alpha 2 and alpha 3 subunits. Co-expression of alpha 4 and beta 1 in Xenopus oocytes resulted in the formation of GABA-gated chloride channels with expected pharmacology, although no benzodiazepine potentiation was observed. Northern analysis indicates that a 4 kb alpha 4 mRNA is expressed in the calf cerebellum, cortex and hippocampus but is barely detectable in the rat brai

Functional properties of recombinant rat GABAA receptors depend upon subunit composition

GABA-gated chloride channels were expressed in human embryonic kidney cells following transfection of cDNAs encoding the alpha 1, beta 2, and gamma 2 subunits of the rat GABAA receptor (GABAR). Functional properties were determined using patch-clamp techniques in the whole-cell and outside-out configurations. Large whole-cell currents were observed in cells expressing the alpha 1 beta 2, alpha 1 gamma 2, and alpha 1 beta 2 gamma 2 subunit combinations. The unique characteristics of GABAR channels consisting of these subunit combinations depended upon the presence or absence of beta 2 and gamma 3 subunits. GABA-activated currents in cells expressing GABARs with the beta 2 subunit desensitized faster and showed greater outward rectification, and the channels had a shorter mean open time than GABARs composed of alpha 1 gamma 2 subunits. When the gamma 2 subunit was present the resulting GABAR channels had a larger conductance. The slope of the concentration-response curve was significantly steeper for GABARs composed of alpha 1 beta 2 gamma 2 subunits compared with GABARs consisting of alpha 1 beta 2 or alpha 1 gamma 2 subunit combinations