Foo Chuan Zhi, Duggan Anne, Bartom Elizabeth T, Tao Litao, García-Añoveros Jaime
Department of Anesthesiology, Northwestern University Feinberg School of Medicine, Chicago, IL, 60611, USA.
Driskill Graduate Program, Northwestern University Feinberg School of Medicine, Chicago, IL, 60611, USA.
J Assoc Res Otolaryngol. 2025 Sep 5. doi: 10.1007/s10162-025-01005-z.
The mammalian cochlea has two types of low abundance and highly specialized inner (IHC) and outer (OHC) mechanosensory hair cells. Their malfunction or death is a common cause of congenital and acquired deafness. IHCs and OHCs exhibit different transcriptomes during development. We wondered how differences in gene expression are regulated at the chromatin level in developing IHCs and OHCs, and whether there were also differences in mRNA splicing between IHCs and OHCs.
We separately collected developing mouse IHCs and OHCs to identify their mRNAs and chromatin states. We examined their transcriptomes by bulk (full coverage) RNA-seq from six biological replicates each to reveal differences in gene expression and in alternative mRNA splicing. We also examined their chromatin conformation by bulk ATAC-seq from two biological replicates each to reveal open vs. closed promoter and enhancer elements. We then compared ATAC-seq with RNA-seq datasets to determine if differential chromatin accessibility can account for differential gene expression. Each biological replicate consists of hair cells pooled from multiple neonatal mice of both sexes.
We found that developing IHCs and OHCs have differentially accessible promoters in many differentially expressed genes. This includes functional genes whose expression is incipient in neonatal hair cells but will be maintained throughout life, and developmental genes which are only expressed transiently. We also found that different mRNA isoforms result from alternative mRNA splicing and transcription start sites. Finally, our data reveals that cochlear hair cells utilize unique promoters and mRNA isoforms absent in other cell types.
Differential transcriptomes between developing hair cell types result from pre- and post-transcriptional mechanisms. The unique promoters and mRNA isoforms in cochlear HCs highlight the importance of elucidating transcriptomes and epigenomes of rare cell types. We provide a comprehensive resource for the identification of promoters and mRNA isoforms of genes expressed by neonatal IHCs or OHCs, which is publicly-accessible for visualization of any gene of interest at https://igvviewer.s3.us-east-2.amazonaws.com/index.html .
哺乳动物的耳蜗有两种数量稀少且高度特化的内毛细胞（IHC）和外毛细胞（OHC）。它们的功能障碍或死亡是先天性和后天性耳聋的常见原因。IHC和OHC在发育过程中表现出不同的转录组。我们想知道在发育中的IHC和OHC中，基因表达的差异在染色质水平上是如何调控的，以及IHC和OHC之间在mRNA剪接方面是否也存在差异。
我们分别收集发育中的小鼠IHC和OHC，以鉴定它们的mRNA和染色质状态。我们通过对每组六个生物学重复样本进行批量（全基因组覆盖）RNA测序来检查它们的转录组，以揭示基因表达和可变mRNA剪接的差异。我们还通过对每组两个生物学重复样本进行批量ATAC测序来检查它们的染色质构象，以揭示启动子和增强子元件的开放与关闭状态。然后，我们将ATAC测序数据集与RNA测序数据集进行比较，以确定染色质可及性差异是否能解释基因表达差异。每个生物学重复样本由来自多只新生雌雄小鼠的毛细胞汇集而成。
我们发现，在许多差异表达基因中，发育中的IHC和OHC具有不同的可及性启动子。这包括在新生毛细胞中开始表达但将在整个生命过程中维持表达的功能基因，以及仅短暂表达的发育基因。我们还发现，可变mRNA剪接和转录起始位点会产生不同的mRNA异构体。最后，我们的数据表明，耳蜗毛细胞利用了其他细胞类型中不存在的独特启动子和mRNA异构体。
发育中的毛细胞类型之间的转录组差异是由转录前和转录后机制导致的。耳蜗毛细胞中独特的启动子和mRNA异构体凸显了阐明稀有细胞类型的转录组和表观基因组的重要性。我们提供了一个全面的资源，用于鉴定新生IHC或OHC表达基因的启动子和mRNA异构体，可通过https://igvviewer.s3.us-east-2.amazonaws.com/index.html公开访问，以可视化任何感兴趣的基因。
