Abstract
It is unclear how loss-of-function germline mutations in the widely-expressed co-chaperone AIP, result in young-onset growth hormone secreting pituitary tumours. The RET receptor, uniquely co-expressed in somatotrophs with PIT1, induces apoptosis when unliganded, while RET supports cell survival when it is bound to its ligand. We demonstrate that at the plasma membrane, AIP is required to form a complex with monomeric-intracellular-RET, caspase-3 and PKCδ resulting in PIT1/CDKN2A-ARF/p53-apoptosis pathway activation. AIP-deficiency blocks RET/caspase-3/PKCδ activation preventing PIT1 accumulation and apoptosis. The presence or lack of the inhibitory effect on RET-induced apoptosis separated pathogenic AIP variants from non-pathogenic ones. We used virogenomics in neonatal rats to demonstrate the effect of mutant AIP protein on the RET apoptotic pathway in vivo. In adult male rats altered AIP induces elevated IGF-1 and gigantism, with pituitary hyperplasia through blocking the RET-apoptotic pathway. In females, pituitary hyperplasia is induced but IGF-1 rise and gigantism are blunted by puberty. Somatotroph adenomas from pituitary-specific Aip-knockout mice overexpress the RET-ligand GDNF, therefore, upregulating the survival pathway. Somatotroph adenomas from patients with or without AIP mutation abundantly express GDNF, but AIP-mutated tissues have less CDKN2A-ARF expression. Our findings explain the tissue-specific mechanism of AIP-induced somatotrophinomas and provide a previously unknown tumorigenic mechanism, opening treatment avenues for AIP-related tumours.
Highlights
The pituitary gland contains five different endocrine cell types with growth hormone (GH)-secreting cells, so-called somatotrophs, contributing to ~50% of the gland
We have explored the action of wild-type human AIP, RETS transfection enhanced more than four times the rate of apoptosis in GH4C1 cells with or without wtAIP co-transfection missense AIP variants identified in pituitary adenoma patients and (Fig. 2B)
We identified that AIP is needed for the normal functioning of the RET/Pit1/ARF/p53-apoptosis pathway (Graphical Abstract, Supplementary Fig. 9)
Summary
The pituitary gland contains five different endocrine cell types with growth hormone (GH)-secreting cells, so-called somatotrophs, contributing to ~50% of the gland. We transfected wt or variant AIP plasmids into GH4C1 cells in the demonstrated that i) overexpression of GDNF is a hallmark of absence or presence of RETS and evaluated apoptosis by Hoechst somatotroph tumours, and ii) lower level ARF expression is a 33258 staining, a system model in which we had extensive previous prognostic marker for resistance to therapy [30]. We had demonstrated that proliferative by any of the AIP variants, except the V49M variant that doubled responses to GHRH varied depending on the cell phenotype, and the basal apoptosis rate (Fig. 2A) This germline variant was this was caused by differences in PIT1 expression [1]. AIP repression (siRNA or knockout) on the RET/PIT1/ARF/p53- expression of wtAIP did not alter GDNF-induced RET activation apoptosis pathway, using well-characterized somatotroph cell lines, and survival.
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