Identification of Cis-Regulatory Sequences Controlling Pollen-Specific Expression of Hydroxyproline-Rich Glycoprotein Genes in Arabidopsis thaliana

Hydroxyproline-rich glycoproteins (HRGPs) are a superfamily of plant cell wall structural proteins that function in various aspects of plant growth and development, including pollen tube growth. We have previously characterized HRGP superfamily into three family members: the hyperglycosylated arabinogalactan-proteins, the moderately glycosylated extensins, and the lightly glycosylated proline-rich proteins. However, the mechanism of pollen-specific HRGP expression remains untouched. To this end, we developed an integrative analysis pipeline combining RNA-seq gene expression and promoter sequences that identified 15 transcriptional cis-regulatory motifs responsible for pollen-specific expression of HRGP in Arabidopsis Thaliana. Specifically, we mined the public RNA-seq datasets and identified 13 pollen-specific HRGP genes. Ensemble motif discovery with various filters identified 15 conserved promoter elements between Thaliana and Lyrata. Known motif analysis revealed pollen related transcription factors of GATA12 and brassinosteroid (BR) signaling pathway regulator BZR1. Lastly, we performed a machine learning regression analysis and demonstrated that the identified 15 motifs well captured the HRGP gene expression in pollen (R=0.61). In conclusion, we performed the integrative analysis as the first-of-its-kind study to identify cis-regulatory motifs in pollen-specific HRGP genes and shed light on its transcriptional regulation in pollen.