Recombinant Expression and Purification of Human Androgen Receptor in a Baculovirus System

Zhu, Zhenxi; Bulgakov, Oleg V.; Scott, Stephanie S.; Dalton, James T.

doi:10.1006/bbrc.2001.5029

Cited by 8 publications

(5 citation statements)

References 36 publications

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“…However, these determinations, although a useful first step, are not definitive because in vitro kinase reactions are often not selective, and mutagenesis can alter the phosphorylations on sites distinct from the ones mutagenized. Ser 308 was directly identified as a phosphorylation site in baculovirus-overexpressed AR using MS [131]. This is the first site identified in living cells either by MS or by in vivo metabolic labelling.…”

Section: Role Of Ar Phosphorylationmentioning

confidence: 95%

Signal transduction in prostate cancer progression

2005

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Prostate cancer is the most frequently diagnosed cancer among men and the second leading cause of male cancer deaths in the United States. When prostate cancer initially presents in the clinic, the tumour is dependent on androgen for growth and, therefore, responsive to the surgical or pharmacological ablation of circulating androgens. However, there is a high rate of treatment failure because the disease often recurs as androgen-independent metastases. Surprisingly, this late-stage androgen-independent prostate cancer almost always retains expression of the AR (androgen receptor), despite the near absence of circulating androgens. Although late-stage prostate cancer is androgen-independent, the AR still seems to play a role in cancer cell growth at this stage of disease. Therefore a key to understanding hormone-independent prostate cancer is to determine the mechanism(s) by which the AR can function even in the absence of physiological levels of circulating androgen. This review will focus on the role of growth factor signalling in prostate cancer progression to androgen independence and thus outline potential molecular areas of intervention to treat prostate cancer progression.

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Section: Role Of Ar Phosphorylationmentioning

confidence: 95%

Signal transduction in prostate cancer progression

2005

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“…However, these determinations, although a useful first step, are not definitive because in vitro kinase reactions are often not selective, and mutagenesis can alter the phosphorylations on sites distinct from the ones mutagenized. Recently, Ser 308 was directly identified as a phosphorylation site in Baculovirus over‐expressed AR using mass spectrometry [Zhu et al, 2001]. This is the first site identified in living cells either by mass spectrometry or by in vivo metabolic labeling.…”

Section: Progression To Hormone “Independence”mentioning

confidence: 99%

Ras signaling in prostate cancer progression

Weber¹,

Gioeli²

2003

J of Cellular Biochemistry

134

110

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When prostate cancer is first detected it generally is dependent on the presence of androgens for growth, and responds to androgen ablation therapies. However, the disease often recurs in a disseminated and apparently androgen independent (AI) form, and in this state is almost invariably fatal. Considerable evidence indicates that the Androgen receptor (AR) continues to be required even in androgen independent (AI) disease. Thus, a key to understanding hormone independent prostate cancer is to determine the mechanism(s) by which the AR can function even in the absence of physiologic levels of androgen. In this article, we argue that growth factors and receptors that utilize Ras family members drive prostate cancer progression to a state of androgen hypersensitivity; and that post-translational modifications (e.g., phosphorylations) of transcriptional cofactors might be responsible for modulating the function of the AR so that it is active even at low concentrations of androgen.

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“…pAR.FLAG expressing the human AR tagged with the FLAG epitope at the COOH terminus is described elsewhere (39). Baculovirus transfer vector pBlueBacHis2A-hAR expressing the full-length human AR is described elsewhere (40).…”

Section: Methodsmentioning

confidence: 99%

“…Bound Tip110-GST fusion protein or GST protein was eluted by 0.4 M reduced glutathione in phosphate-buffered saline (PBS), and the glutathione was removed by extensive dialysis against PBS. For the full-length recombinant AR protein, preparation of baculovirus stock, infection of SF9 cells, and AR purification were performed exactly as previously described (40). The purity of GST, Tip110-GST, and AR protein was determined to be higher than 95% by SDS-PAGE and Coomassie staining.…”

Section: Methodsmentioning

confidence: 99%