PROJECT SUMMARY
The formation of comedones, referred to as comedogenesis, describes the production of keratinized plug that
occludes the opening of the hair follicle. Comedones are often observed in common cutaneous diseases like
acne vulgaris, with up to 85% of individuals experiencing comedonal lesions in their lifetime. Comedones are
also seen in rare, severe diseases such as nevus comedonicus (NC), a disease involving comedones that
progress into painful, inflammatory cysts. We have shown that NC is caused by somatic mutations in the NEK9
gene. Remarkably, whole exome sequencing (WES) of a common comedonal disorder, the epidermal inclusion
cyst, revealed identical mutations in NEK9, suggesting NEK9 mutations may drive comedogenesis across
different manifestations. While NEK9 has been well studied in the context of mitosis and cell cycle progression,
sporadic associations of NEK9 with primary cilia are also seen in the literature. The primary cilium, an immotile
finger-like projection from the cell with critical functions in signal transduction, houses many of the signaling
pathways related to proper development and maintenance of hair follicles. Because comedones arise from
follicular structures, these mutations in NEK9 provide an unmatched opportunity to elucidate the genetic and
biologic mechanisms underlying comedogenesis. To accomplish this, we have generated in vitro and in vivo
models of NEK9 mutations identified in NC and EIC. Our preliminary data show NEK9 mutations result in shorter
cilia and less ciliation in vitro. Hedgehog signaling, a key follicular signaling pathway that acts through the primary
cilium, is diminished in NEK9 mutant cells. Our immunofluorescence studies show the activated form of the
NEK9 protein localizes to the base of the cilium, a new discovery in NEK9 biology. We have previously shown
this activation signal to be increased across the NEK9 mutations. Together, these preliminary data support ciliary
dysfunction to be a result of NEK9 mutations. To study the involvement of NEK9 and cilia in comedogenesis in
vivo, we have generated a transgenic mouse using a tamoxifen-dependent, basal keratinocyte specific promoter
(K14-CreERT) to drive transcription of the NEK9 mutation most commonly reported in NC. We will time transgene
induction with follicular morphogenesis onset and perform immunofluorescence and advanced microscopy
methods to interrogate defects in canonical follicular development. We will continue to obtain NC and EIC tissue
from our institution and perform WES to identify new candidate variants that may contribute to comedogenesis.
We will generate stratified skin equivalent models to continually test the newly identified mutations for ciliary
phenotypes. Lastly, we will use the known biology of NEK9 to systematically test downstream effectors for
possible mechanisms of ciliary dysfunction. This training proposal represents an integrated scientific approach
and new learning experiences that harness techniques in cell biology, computational genetics, and pathology to
yield novel insights into the mechanisms of comedogenesis.
