FGFR2 mutations in bent bone dysplasia syndrome activate nucleolar stress and perturb cell fate determination

Abstract

Fibroblast Growth Factor (FGF) signaling promotes self-renewal in progenitor cells by encouraging proliferation and inhibiting cellular senescence. Yet, these beneficial effects can be hijacked by disease-causing mutations in FGF receptor (FGFR) during embryogenesis. By studying dominant FGFR2 mutations that are germline in bent bone dysplasia syndrome (BBDS), we reveal a mechanistic connection between FGFR2, ribosome biogenesis, and cellular stress that links cell fate determination to disease pathology. We previously showed that FGFR2 mutations in BBDS, which amplify nucleolar targeting of FGFR2, activate ribosomal DNA (rDNA) transcription and delay differentiation in osteoprogenitor cells and patient-derived bone. Here we find that the BBDS mutations augment the ability of FGFR2 to recruit histone-remodeling factors that epigenetically activate transcriptionally silent rDNA. Nucleolar morphology is controlled by chromatin structure, and the high levels of euchromatic rDNA induced by the BBDS mutations direct nucleolar disorganization, alter ribosome biogenesis, and activate the Rpl11-Mdm2-p53 nucleolar stress response pathway. Inhibition of p53 in cells expressing the FGFR2 mutations in BBDS rescues delayed osteoblast differentiation, suggesting that p53 activation is an essential pathogenic factor in, and potential therapeutic target for, BBDS. This work establishes rDNA as developmentally regulated loci that receive direct input from FGF signaling to balance self-renewal and cell fate determination.

FGF/FGFR signaling in health and disease

“…FGFR2 mutations (FGFR2 M391R and FGFR2 Y381D ) located in the TMD can reduce plasma membrane levels of FGFR2, and amplify its nuclear and nucleolar presence in growth plate chondrocytes derived from patients with skeletal disorder bent bone dysplasia syndrome (BBDS). 95,96 Interestingly, posttranslational modifications, such as glycosylation, also contribute to the nuclear localization of FGFRs. In the skeletal disorder Crouzon syndrome, the FGFR2 mutation (FGFR2 C278F ) leads to incomplete FGFR2 glycosylation, blocks its membrane localization, and induces the perinuclear accumulation of receptor.…”

“…189 Further studies on BBDS resulting from FGFR2 mutations revealed that nuclear FGFR2 regulates the developing limb, musculoskeletal integration, and cell fate determination. 96,192 Targeted disruption of FGFR2IIIc in mice leads to narrowed proliferative and hypertrophic zones in growth plate, and disturbed ossification with downregulation of IHH, PTHRP, and RUNX2. 17 Yu et al 193 found that conditional deletion of FGFR2 in mesenchyme can lead to skeletal dwarfism and decreased bone mineral density with dramatically disturbed proliferation of osteoprogenitors and anabolic function of mature osteoblasts in mice.…”

Nuclear FGFR2 regulates musculoskeletal integration within the developing limb

“…Previously, we showed that the BBDS mutations enhance nuclear and nucleolar localization of FGFR2 in patient‐derived chondrocytes and osteoprogenitor cells (Merrill et al, ; Neben et al, , ). In ATDC5 prechondrocytes, immunofluorescent localization of FLAG‐tagged FGFR2 M391R and FGFR2 Y381D and the nucleolar marker Polr1D confirmed that these mutations enhanced receptor localization to the nucleus and nucleolus of chondrogenic cells (Fig.…”

“…While the role of FGFRs as transmembrane signal transducers is well-defined, it is unclear how these receptors produce a transcriptional signature distinct from other receptor tyrosine kinases that use the same downstream signaling cascades. Our lab and others have shown that FGFRs and their ligands act in the nucleus to regulate gene expression (Reilly et al, 2004;Sorensen et al, 2006;Stachowiak et al, 2007Stachowiak et al, , 2015Coleman et al, 2014;Neben et al, 2014Neben et al, , 2017. Although nuclear FGFR signaling could provide insight into FGF signaling specificity in development and disease, little is known about the significance of this activity in vivo.…”

“…We showed previously that the BBDS mutations FGFR2 M391R and FGFR2 Y381D are located in the transmembrane domain of FGFR2 and enhance receptor localization to the nucleus in skeletal progenitor cells (Merrill et al, 2012;Neben et al, 2014Neben et al, , 2017. While we are beginning to understand the molecular consequences of the BBDS mutations on nuclear FGFR2 signaling, it remains unclear how altered localization of FGFR2 affects skeletal development in vivo.…”