Identification of an alternative ligand-binding pocket in the nuclear vitamin D receptor and its functional importance in 1α,25(OH) ₂ -vitamin D ₃ signaling

Abstract

T he steroid hormone 1␣,25(OH) 2 -vitamin D 3 (1,25D) (Fig. 1A), other steroid hormones, retinoids, and thyroid hormones form the family of ligands for the nuclear receptor (NR) superfamily (1), the members of which produce genomic responses through selective interaction of the liganded receptor with promoters of appropriate genes and basal transcription machinery. Many of these hormones also activate rapid, nongenomic (NG), cellular signaling cascades (2, 3) (except retinoids) that range from activation of ion channels (4, 5) to promoting kinase and other cytosolic signaling cascades (6-9). Defining the structure-function requirements for 1,25D and 17␤-estradiol (E 2 ) rapid actions has been aided by the synthesis of analogs that are NG agonists like 1␣,25(OH) 2 -lumisterol (JN) (ref. 10 and Fig. 1 A) and 4-estren-3␣,17␤-diol (EST) (11) or antagonists like 1␤,25(OH) 2 -vitamin D 3 (HL) (ref. 12 and Fig. 1 A), but that are only weak genomic transactivators (13). Thus, important structural attributes of the sterol dictate its agonistic properties and subsequent genomic vs. NG signaling profile (14, 15).When 1,25D rapid signaling cascades were first discovered, it was hypothesized that the observed activities were propagated by a novel membrane protein(s) (16), because analogs JN and HL did not compete well with [ 3 H]1,25D for binding to the nuclear vitamin D receptor (VDR) (17). Recently, in studies using a VDR knockout (KO) mouse (18) and a naturally occurring human VDR mutation (19), 1,25D-mediated rapid responses were shown to require a functional VDR. Both the VDR and estrogen receptor (ER) have been found localized to the plasma membrane in caveolae (7, 20); therefore, it has been proposed that the VDR and ER propagate some NG signaling (6,9,18,21,22). However, given the poor affinity of JN and HL for the VDR, it is difficult to understand how these sterols can facilitate their activities through the VDR.Results obtained from the modeling (INSIGHT 2000.1) of JN, 1,25D, and HL in the VDR ligand-binding domain (LBD) showed that the VDR could possibly accept and form favorable nonbonding interactions with vitamin D sterols in a distinct ligand-binding pocket [an alternative ligand-binding pocket (A pocket)] from the genomic pocket (G pocket) that was previously defined by x-ray crystallography (23, 24). Our proposed A-pocket accepts ligands that differ in shape from those in the classical G pocket (3,23,24).The data from these models has led to the proposal that the VDR can function as a rapid response receptor through a conformational ensemble mechanism (3, 25) whereby the flexible 1,25D steroid hormone samples an ensemble of energetically similar protein conformations (26). In addition, the ensemble model and existence of an A pocket may provide an explanation for the observed sex-nonspecific, nongenotropic signaling through the ER␣ receptor by EST (3, 9, 11). The physiological relevance of the ensemble model and functional importance of an A pocket within the VDR is further substantiated by applying the m...

1α,25(OH)2-Vitamin D and a Nongenomic Vitamin D Analogue Inhibit Ultraviolet Radiation–Induced Skin Carcinogenesis

“…These results complement those of Ellison and colleagues, who showed that VDR-knockout mice are much more susceptible to photocarcinogenesis (44). The VDR has been shown to be essential for nongenomic effects of vitamin D compounds (9,10,13). In VDR-knockout mice, thymine dimer repair was significantly reduced (44 (44) is that photoprotection in mice seems to be mediated through a nongenomic pathway, which is still likely to involve the VDR but which may be activated by compounds other than the classical hormone (10).…”

“…Thus, the same 1,25(OH) 2 D 3 nuclear receptor (VDR) has been found in the plasma membrane caveolae of target cells where it is engaged in mediating nongenomic responses (9,10). The VDR ligand 1,25(OH) 2 D 3 is highly conformationally flexible, and it has been shown that one shape of 1,25(OH) 2 D 3 is preferred for genomic responses whereas another shape is preferred by the VDR for nongenomic responses (11)(12)(13). The 6-s-cis conformationally restricted analogue 1a,25(OH) 2 -lumisterol (JN) shows little ability to increase transcription of the osteocalcin gene in bone cells compared with 1,25(OH) 2 D 3 (14) but generates nongenomic actions in pancreatic b cells and endothelial cells (15,16).…”

New insights into Vitamin D sterol-VDR proteolysis, allostery, structure–function from the perspective of a conformational ensemble model

“…1C) [25]. The VDR A-pocket was discovered using in silico techniques and has been linked to shape and stereospecific induction of nongenomic cellular responses [26], as well as the unique structure-function results obtained for some A-ring and side-chain derivatives of 1,25D [27,28]. An important assumption made in these works is that the VDR A-pocket represents a kinetically favored ligand binding pocket (LBP), while x-ray results clearly indicate the VDR G-pocket is the thermodynamically favored LBP.…”

Vitamin D receptor distribution in intestines of domesticated sheep Ovis ammon f. aries

“…These include interaction of calcitriol with a membrane receptor, either a novel receptor or the known vitamin D receptor, thereby activating cell signalling pathways (Nemere and Campbell, 2000;Holick, 2003;Evans et al, 2004;Fleet, 2004;Mizwicki et al, 2004;Dusso et al, 2005;Hendy et al, 2006). There is growing direct evidence that the traditional vitamin D receptor may also have a unique, nontranscriptional role in plasma membrane initiated signalling (Fleet, 2004 (Berger et al, 1988;Sandgren et al, 1991;Rougui et al, 1998;Sheinin et al, 2000;Chatterjee, 2001;Holick, 2003;Lee et al, 2003).…”