Specification of Stem Cells and Niche During Hair Follicle Morphogenesis

Adult stem cell (SC) behavior is tightly coordinated by the signals received from the “niche” - the microenvironment that the SCs reside in. Little is known about the role of the niche in SC specification during organ morphogenesis. In particular, the question of whether the niche exists prior to SC specification or whether it is recruited after SC establishment in a developing tissue remains largely unanswered. In addition, the signals responsible for the specification and regulation of SCs during morphogenesis remain unexplored. To answer these questions, I focused my analysis on the earliest stages of hair follicle (HF) morphogenesis. Using immunofluorescence and live imaging, I found that in developing HFs, basal cell divisions are asymmetric and perpendicular to the basement membrane. These divisions result in differential levels of WNT signaling in the daughter cells, with basal cells remaining WNThigh, and suprabasal cells becoming WNTlow. Using in utero lentiviral transduction and genetic mouse models, I created mosaic epidermis with gain- or loss-of-function for WNT signaling to demonstrate that juxtaposition of WNTlow and WNThigh cells is sufficient to confer SOX9+ cell fate to the WNTlow cells. This suggested that the perpendicular asymmetric divisions that I observed in the developing HFs produce the WNT gradient, necessary for the establishment of SOX9+ cells. To further investigate the mechanism behind SOX9+ cell specification, I investigated the signaling patterns of SHH, previously suggested to regulate Sox9 expression. Interestingly, while Shh was expressed exclusively by the WNThigh basal cells, SHH signaling was primarily detected in the suprabasal SOX9+ cells. By inducing the expression of lentivirus-delivered Shh at different stages in morphogenesis, I found that the levels of WNT signaling dictate the responsiveness to SHH. When WNT signaling is low or moderate, cells respond to SHH, resulting in the inhibition of WNT signaling. However, WNThigh cells are resistant to SHH signaling. Thus, in the suprabasal SOX9+ daughters, SHH acts to repress WNT signaling, further boosting the levels of SOX9, while due to high levels of WNT signaling in the basal daughters, they are unable to respond to SHH and remain SOX9-negative. Finally, using Shh-CreER lineage-tracing, I demonstrated that the earliest asymmetric cell divisions of the WNThigh, Shh+ cells produce SOX9+ cells that eventually contribute to the adult stem cell pool. Interestingly, SOX9+ cells are produced only during the early asymmetric cell divisions, while the same Shh+ cells later give rise to various differentiated lineages of the developing HF. Thus, in developing HFs, asymmetric cell divisions produce a WNTlow SOX9+ SC daughter and a WNThigh “niche” cell daughter that produces SHH, necessary to suppress WNT signaling and expand the SOX9+ SCs

A large peptidome dataset improves HLA class I epitope prediction across most of the human population

Published in final edited form as: Nat Biotechnol. 2020 February ; 38(2): 199–209. doi:10.1038/s41587-019-0322-9.Prediction of HLA epitopes is important for the development of cancer immunotherapies and vaccines. However, current prediction algorithms have limited predictive power, in part because they were not trained on high-quality epitope datasets covering a broad range of HLA alleles. To enable prediction of endogenous HLA class I-associated peptides across a large fraction of the human population, we used mass spectrometry to profile >185,000 peptides eluted from 95 HLA-A, -B, -C and -G mono-allelic cell lines. We identified canonical peptide motifs per HLA allele, unique and shared binding submotifs across alleles and distinct motifs associated with different peptide lengths. By integrating these data with transcript abundance and peptide processing, we developed HLAthena, providing allele-and-length-specific and pan-allele-pan-length prediction models for endogenous peptide presentation. These models predicted endogenous HLA class I-associated ligands with 1.5-fold improvement in positive predictive value compared with existing tools and correctly identified >75% of HLA-bound peptides that were observed experimentally in 11 patient-derived tumor cell lines.P01 CA229092 - NCI NIH HHS; P50 CA101942 - NCI NIH HHS; T32 HG002295 - NHGRI NIH HHS; T32 CA009172 - NCI NIH HHS; U24 CA224331 - NCI NIH HHS; R21 CA216772 - NCI NIH HHS; R01 CA155010 - NCI NIH HHS; U01 CA214125 - NCI NIH HHS; T32 CA207021 - NCI NIH HHS; R01 HL103532 - NHLBI NIH HHS; U24 CA210986 - NCI NIH HHSAccepted manuscrip