Parafibromin, product of the hyperparathyroidism-jaw tumor syndrome gene HRPT2, regulates cyclin D1/PRAD1 expression

Abstract

Parafibromin is the 531-amino-acid protein product encoded by HRPT2, a putative tumor suppressor gene recently implicated in the autosomal dominant hyperparathyroidism-jaw tumor familial cancer syndrome, sporadic parathyroid cancer, and a minority of families with isolated hyperparathyroidism. Parafibromin contains no identified functional domains but bears sequence homology to Cdc73p, a budding yeast protein component of the RNA polymerase II-associated Paf1 complex. This study addressed the expression and functional properties of human parafibromin. A survey of human and mouse tissues analysed with polyclonal antibodies to parafibromin showed specific immunoreactivity in adrenal and parathyroid glands, kidney, heart, and skeletal muscle. Subcellular fractionation and laser confocal microscopy of normal human parathyroid gland demonstrated expression of parafibromin in both the cytoplasmic and nuclear compartments. Parafibromin was expressed in four parathyroid adenomas but was absent from two parathyroid carcinomas. Transient overexpression of wild-type parafibromin, but not its Leu64Pro missense mutant implicated in parathyroid cancer and familial isolated hyperparathyroidism, inhibited cell proliferation, and blocked expression of cyclin D1, a key cell cycle regulator previously implicated in parathyroid neoplasia. These results demonstrate that human parafibromin is a nucleocytoplasmic protein with functions consistent with its postulated role as a tumor suppressor protein.

Sporadic human renal tumors display frequent allelic imbalances and novel mutations of the HRPT2 gene

“…Concomittantly, HA-tagged species of wild-type parafibromin (lane 3), the R222X mutant lacking the C-terminal PAF1-binding domain (lane 4) and the newly identified K34Q mutant derivative (lane 5) were efficiently produced. Consistent with previous findings demonstrating that parafibromin overexpression is correlated with decreased cyclin D1 expression (Woodard et al, 2005), we observed that downregulation of endogenous parafibromin caused an increase in cyclin D1 protein levels (compare lane 2 and 1). Re-expression of HA-tagged wild-type parafibromin reverted this effect (lane 3).…”

“…Mutation in parafibromin have been recently linked to the control of cyclin D1 expression (Woodard et al, 2005). Thus, we next determined whether the K34Q parafibromin mutant is defective in this regard.…”

“…Finally, ectopic expression of parafibromin has been shown to inhibit the expression of the cell cycle regulatory protein cyclin D1. Interestingly, a point mutant derivative of it, L64P, failed to inhibit cyclin D1 expression (Woodard et al, 2005). The latter provides an intriguing link between normal parafibromin function, cell cycle control and parathyroid cancer suppression.…”

Parafibromin is a nuclear protein with a functional monopartite nuclear localization signal

“…Furthermore, transient over-expression of parafibromin inhibits cyclin D1 expression whereas downregulation of parafibromin using small interfering RNA (siRNA) leads to premature entry of cells into S phase (Woodard et al, 2004;Yart et al, 2005). In addition, parafibromin is associated with a Set-1-like histone methyltransferase (HMTase) complex that methylates H3 on lysine 4, and thereby initiates transcription by chromatin modification and activation of RNA polymerase II (Rozenblatt-Rosen et al, 2005;Yart et al, 2005).…”

“…Thus, as part of this protein complex, parafibromin may regulate posttranscriptional events and histone modification. In keeping with such roles parafibromin has been reported to be a nuclear protein by two studies (Tan et al, 2004;Rozenblatt-Rosen et al, 2005), although one other study has reported that parafibromin may also be in the cytoplasm (Woodard et al, 2004).…”