Biology subjects

McCurdy, B. L.

Publications and source records attributed to McCurdy, B. L..

6 recordsLinked to original sources

bioRxiv2026

Aberrant PCM1 accumulation in Trisomy 21 mislocalizes E3 ligases, delaying primary ciliogenesis

Primary cilia are microtubule-based extracellular signaling structures essential for development and tissue homeostasis, and their defects can cause ciliopathies. Trisomy 21, the cause of Down syndrome, also disrupts cilia formation and function. Here we show that Pericentrin (PCNT), a chromosome 21 resident gene whose protein is elevated in Trisomy 21, impairs primary ciliogenesis by delaying mother centriole uncapping. Elevated PCNT forms pericentrosomal assemblies that promote the accumulation of PCM1 in the pericentrosomal compartment. PCM1 binds and localizes the CP110 E3 ubiquitin ligases HERC2 and MIB1 to the centrosome, so mislocalizing PCM1 depletes these ligases from the centrosome, lowering CP110 ubiquitination and delaying uncapping. Reducing PCNT rescues these defects, whereas elevating PCM1 phenocopies them, establishing PCM1 accumulation as a critical mediator of uncapping. These findings reveal how a dosage-sensitive chromosome 21 gene disrupts ciliogenesis and show that centrosome function depends not only on PCM protein composition but on the spatial partitioning of proteins between the centrosome and pericentrosomal compartments.

cell biology↗

bioRxiv2026

Trisomy 21 alters ciliary localization of Sonic Hedgehog signaling proteins

Trisomy 21 (T21), the cause of Down syndrome, impairs cilia-dependent Sonic Hedgehog (SHH) signaling. We investigated how T21 affects SHH signaling at the primary cilium and found that Smoothened (SMO) protein - the main effector of SHH signaling - is diminished in T21 primary cilia. Specifically, T21 cells exhibit both decreased SMO localization to cilia and reduced ciliary SMO phosphorylation (pSMO). Our findings suggest that reduced ciliary SMO in T21 results from defective entry into or maintenance in the cilium rather than impaired transport to centrosomes. Moreover, unlike the ciliogenesis defects observed in T21, the SMO defect does not appear to be associated with increased PCNT protein abundance. We also found that GPR161 (a negative SHH regulator), exhibits increased ciliary localization in T21. Notably, prolonged serum depletion to promote primary cilia maturation equalizes ciliary levels of SMO and GPR161 between T21 cells and D21 controls. We propose that delayed primary cilia maturation contributes to defective SHH signaling in T21. SUMMARYT21 impedes primary cilia assembly and disrupts SHH signaling by diminishing ciliary SMO and increasing GPR161, but extended ciliary maturation equalizes SHH protein localization between D21 and T21.

cell biology↗

bioRxiv2025

Splice-switching antisense oligonucleotides correct cryptic exon inclusion and restore SDCCAG8 protein in Bardet-Biedl Syndrome

Bardet-Biedl Syndrome (BBS) is a ciliopathy often associated with progressive blindness and obesity. A patient presenting with BBS was discovered to have two mutations within 55bp of each other in intron 7 of SDCCAG8 (BBS16). One of the biallelic mutations, c.740+356C>T, causes inclusion of cryptic exon(s) containing premature termination codons, while c.740+301G>A has not been characterized. We hypothesized that antisense oligonucleotides (ASOs) complementary to the patients mutations or to the cryptic exon splice sites would correct the splicing of SDCCAG8 between exons 7 and 8 to prevent cryptic exon inclusion and restore SDCCAG8 expression. We systematically screened 20nt-long ASOs tiled across each mutation and ASOs targeting the 3' splice sites of the cryptic exons in patient-derived fibroblasts, using RT-PCR assays to assess exon 7 and 8 splicing. We identified one ASO for each mutation and a cryptic exon-targeting ASO that restored the splicing pattern to that observed in an unaffected cell line. Lead ASOs were further investigated through RT-PCR, RNA sequencing, and western blotting to confirm ASO-mediated restoration of wild-type transcript and protein. Notably, ASO 20, which targets the cryptic exon 7a/7a' splice site rather than patient-specific mutations, achieved the greatest rescue effect, increasing exon 7-8 splicing from 0% to an average of 26% and restoring SDCCAG8 protein from undetectable levels to approximately 40% of wild-type expression. This mutation-agnostic approach could benefit multiple patients with cryptic exon inclusion in this region of SDCCAG8, expanding therapeutic impact beyond traditional N-of-1 ASO strategies. These findings establish a molecular foundation for clinical development of ASO therapy for BBS caused by SDCCAG8 splicing defects.

genetics↗

bioRxiv2023

TTLL12 is required for primary ciliary axoneme formation in polarized epithelial cells

The primary cilium is a critical sensory organelle that is built of axonemal microtubules ensheathed by a ciliary membrane. In polarized epithelial cells, primary cilia reside on the apical surface and must extend these microtubules directly into the extracellular space and remain a stable structure. However, the factors regulating cross-talk between ciliation and cell polarization, as well as, axonemal microtubule growth and stabilization in polarized epithelia are not fully understood. In this study, we find TTLL12, a previously uncharacterized member of the Tubulin Tyrosine Ligase-Like (TTLL) family, localizes to the base of primary cilia and is required for cilia formation in polarized renal epithelial cells. We also show that TTLL12 directly binds to the /{beta}-tubulin heterodimer in vitro and regulates microtubule dynamics, stability, and post-translational modifications (PTMs). While all other TTLLs catalyze the addition of glutamate or glycine to microtubule C-terminal tails, TTLL12 uniquely affects tubulin PTMs by promoting both microtubule lysine acetylation and arginine methylation. Together, this work identifies a novel microtubule regulator and provides insight into the requirements for apical extracellular axoneme formation.

cell biology↗

bioRxiv2021

Trisomy 21 induces pericentrosomal crowding disrupting early stages of primary ciliogenesis and mouse cerebellar development

Primary cilia are signaling organelles essential for development and homeostasis. Loss of primary cilia is lethal, and decreased or defective cilia cause multisystemic conditions called ciliopathies. Down syndrome shares clinical overlap with ciliopathies. We previously showed that trisomy 21 diminishes primary cilia formation and function due to elevated Pericentrin, a centrosome protein encoded on chromosome 21. Pericentrin is mislocalized, creating aggregates that disrupt pericentrosomal trafficking and microtubule organization. Here, we examine the cilia-related molecules and pathways disrupted in trisomy 21 and their in vivo phenotypic relevance. Utilizing ciliogenesis time course experiments, we reveal how Pericentrin, microtubule networks, and components of ciliary vesicles are reorganized for ciliogenesis in euploid cells. Early in ciliogenesis, chromosome 21 polyploidy results in elevated Pericentrin and microtubule networks away from the centrosome that ensnare MyosinVA and EHD1, blocking mother centriole uncapping that is essential for ciliogenesis. Ciliated trisomy 21 cells have persistent trafficking defects that reduce transition zone protein localization, which is critical for Sonic hedgehog signaling. Sonic hedgehog signaling is decreased and anticorrelates with Pericentrin levels in trisomy 21 primary mouse embryonic fibroblasts. Finally, we observe decreased ciliation in vivo. A mouse model of Down syndrome with elevated Pericentrin has fewer primary cilia in cerebellar granule neuron progenitors and thinner external granular layers. Our work reveals that elevated Pericentrin in trisomy 21 disrupts multiple early steps of ciliogenesis and creates persistent trafficking defects in ciliated cells. This pericentrosomal crowding results in signaling defects consistent with the neurological deficits found in individuals with Down syndrome.

cell biology↗

bioRxiv2021

Trisomy 21 increases microtubules and disrupts centriolar satellite localization

Trisomy 21, the cause of Down syndrome, causes a 0.5-fold protein increase of the chromosome 21-resident gene Pericentrin (PCNT) and reduces primary cilia formation and signaling. Here we investigate the mechanisms by which PCNT imbalances disrupt cilia. Using isogenic RPE-1 cells with increased chromosome 21 dosage, we find PCNT protein accumulates around the centrosome as a pericentrosomal cluster of enlarged cytoplasmic puncta that localize along and at MT ends. Cytoplasmic PCNT puncta impact the intracellular MT trafficking network required for primary cilia, as the PCNT puncta sequester cargo peripheral to centrosomes in what we call pericentrosomal crowding. The centriolar satellite proteins, PCM1, CEP131 and CEP290, important for ciliogenesis, accumulate at sites of enlarged PCNT puncta in trisomy 21 cells. Reducing PCNT when chromosome 21 ploidy is elevated is sufficient to decrease PCNT puncta, reestablish a normal density of MTs around the centrosome, restore ciliogenesis to wild type levels and decrease pericentrosomal crowding. A transient reduction in MTs also decreases pericentrosomal crowding and partially rescues ciliogenesis in trisomy 21 cells, indicating that increased PCNT leads to defects in the microtubule network deleterious to normal centriolar satellite distribution. We propose that chromosome 21 aneuploidy disrupts MT-dependent intracellular trafficking required for primary cilia. TOCMcCurdy et al explore why elevated Pericentrin in trisomy 21 negatively impacts primary cilia formation and signaling. They find that elevated Pericentrin produces more pericentrosomal puncta that associates with and increases microtubules. Changes to Pericentrin and microtubules mislocalizes centriolar satellites in a pericentrosomal crowd.

cell biology↗