TY - JOUR
T1 - Analysis of FGF20-regulated genes in organ of Corti progenitors by translating ribosome affinity purification
AU - Yang, Lu M.
AU - Stout, Lisa
AU - Rauchman, Michael
AU - Ornitz, David M.
N1 - Funding Information:
We thank J. Dougherty and his laboratory for their help with the TRAP technique, B. Zhang (Department of Developmental Biology), and T. Sinnwell and M. Heinz (Genome Technology Access Center in the Department of Genetics) for their help with RNA sequencing and analysis. We thank L. Robbins for generating the , , and plasmids used for making in situ probes. We also thank C. Ferguson for supplying us with mice. This work was funded by the Department of Developmental Biology at Washington University, NIH/National Institute on Deafness and Other Communication Disorders grant DC017042 (DMO), the Washington University Institute of Clinical and Translational Sciences which is, in part, supported by the NIH/National Center for Advancing Translational Sciences, CTSA grant UL1TR002345 (JIT471 to DMO), March of Dimes grant 6‐FY13‐127 (MR), and the Rare Disease Foundation/BC Children's Hospital Foundation. GTAC is partially supported by NCI Cancer Center Support Grant P30 CA91842 to the Siteman Cancer Center and by ICTS/CTSA Grant UL1TR000448 from the National Center for Research Resources (NCRR). HOPE Center Alafi Neuroimaging Laboratory is supported by NCRR grant 1S10RR027552, the Auditory and Vestibular Microscopy and Digital Imaging Core is supported by NIH grant P30 DC004665. Sall1 Sall2 Sall3 Cdc20 flox
Funding Information:
March of Dimes Foundation, Grant/Award Number: 6‐FY13‐127; National Center for Advancing Translational Sciences, Grant/Award Number: UL1TR002345; National Institute on Deafness and Other Communication Disorders, Grant/Award Number: DC017042; Rare Disease Foundation/BC Children's Hospital Foundation; NCA Cancer Center, Grant/Award Number: P30 CA91842; ICTS/CTSA, Grant/Award Number: UL1TR000448; National Center for Research Resources (NCRR), Grant/Award Number: 1S10RR027552; Washington University Institute of Clinical and Translational Sciences; NIH, Grant/Award Number: P30 DC004665; CTSA, Grant/Award Number: JIT471 Funding information P
Funding Information:
We thank J. Dougherty and his laboratory for their help with the TRAP technique, B. Zhang (Department of Developmental Biology), and T. Sinnwell and M. Heinz (Genome Technology Access Center in the Department of Genetics) for their help with RNA sequencing and analysis. We thank L. Robbins for generating the Sall1, Sall2, and Sall3 plasmids used for making in situ probes. We also thank C. Ferguson for supplying us with Cdc20flox mice. This work was funded by the Department of Developmental Biology at Washington University, NIH/National Institute on Deafness and Other Communication Disorders grant DC017042 (DMO), the Washington University Institute of Clinical and Translational Sciences which is, in part, supported by the NIH/National Center for Advancing Translational Sciences, CTSA grant UL1TR002345 (JIT471 to DMO), March of Dimes grant 6-FY13-127 (MR), and the Rare Disease Foundation/BC Children's Hospital Foundation. GTAC is partially supported by NCI Cancer Center Support Grant P30 CA91842 to the Siteman Cancer Center and by ICTS/CTSA Grant UL1TR000448 from the National Center for Research Resources (NCRR). HOPE Center Alafi Neuroimaging Laboratory is supported by NCRR grant 1S10RR027552, the Auditory and Vestibular Microscopy and Digital Imaging Core is supported by NIH grant P30 DC004665.
Publisher Copyright:
© 2020 Wiley Periodicals LLC
PY - 2020/10/1
Y1 - 2020/10/1
N2 - Background: Understanding the mechanisms that regulate hair cell (HC) differentiation in the organ of Corti (OC) is essential to designing genetic therapies for hearing loss due to HC loss or damage. We have previously identified Fibroblast Growth Factor 20 (FGF20) as having a key role in HC and supporting cell differentiation in the mouse OC. To investigate the genetic landscape regulated by FGF20 signaling in OC progenitors, we employ Translating Ribosome Affinity Purification combined with Next Generation RNA Sequencing (TRAPseq) in the Fgf20 lineage. Results: We show that TRAPseq targeting OC progenitors effectively enriched for RNA from this rare cell population. TRAPseq identified differentially expressed genes (DEGs) downstream of FGF20, including Etv4, Etv5, Etv1, Dusp6, Hey1, Hey2, Heyl, Tectb, Fat3, Cpxm2, Sall1, Sall3, and cell cycle regulators such as Cdc20. Analysis of Cdc20 conditional-null mice identified decreased cochlea length, while analysis of Sall1-null and Sall1-ΔZn2-10 mice, which harbor a mutation that causes Townes-Brocks syndrome, identified a decrease in outer hair cell number. Conclusions: We present two datasets: genes with enriched expression in OC progenitors, and DEGs downstream of FGF20 in the embryonic day 14.5 cochlea. We validate select DEGs via in situ hybridization and in vivo functional studies in mice.
AB - Background: Understanding the mechanisms that regulate hair cell (HC) differentiation in the organ of Corti (OC) is essential to designing genetic therapies for hearing loss due to HC loss or damage. We have previously identified Fibroblast Growth Factor 20 (FGF20) as having a key role in HC and supporting cell differentiation in the mouse OC. To investigate the genetic landscape regulated by FGF20 signaling in OC progenitors, we employ Translating Ribosome Affinity Purification combined with Next Generation RNA Sequencing (TRAPseq) in the Fgf20 lineage. Results: We show that TRAPseq targeting OC progenitors effectively enriched for RNA from this rare cell population. TRAPseq identified differentially expressed genes (DEGs) downstream of FGF20, including Etv4, Etv5, Etv1, Dusp6, Hey1, Hey2, Heyl, Tectb, Fat3, Cpxm2, Sall1, Sall3, and cell cycle regulators such as Cdc20. Analysis of Cdc20 conditional-null mice identified decreased cochlea length, while analysis of Sall1-null and Sall1-ΔZn2-10 mice, which harbor a mutation that causes Townes-Brocks syndrome, identified a decrease in outer hair cell number. Conclusions: We present two datasets: genes with enriched expression in OC progenitors, and DEGs downstream of FGF20 in the embryonic day 14.5 cochlea. We validate select DEGs via in situ hybridization and in vivo functional studies in mice.
KW - RNAseq
KW - SALL1
KW - Townes-Brocks syndrome
KW - cochlea
KW - hair cell
KW - hearing loss
UR - http://www.scopus.com/inward/record.url?scp=85087738997&partnerID=8YFLogxK
U2 - 10.1002/dvdy.211
DO - 10.1002/dvdy.211
M3 - Article
C2 - 32492250
AN - SCOPUS:85087738997
VL - 249
SP - 1217
EP - 1242
JO - Developmental Dynamics
JF - Developmental Dynamics
SN - 1058-8388
IS - 10
ER -