Comprehensive transcriptome analysis of phytohormone biosynthesis and signaling genes in microspore/pollen and tapetum of rice

Abstract

To investigate the involvement of phytohormones during rice microspore/pollen (MS/POL) development, endogenous levels of IAA, gibberellins (GAs), cytokinins (CKs) and abscisic acid (ABA) in the mature anther were analyzed. We also analyzed the global expression profiles of genes related to seven phytohormones, namely auxin, GAs, CKs, brassinosteroids, ethylene, ABA and jasmonic acids, in MS/POL and tapetum (TAP) using a 44K microarray combined with a laser microdissection technique (LM-array analysis). IAA and GA(4) accumulated in a much higher amount in the mature anther compared with the other tissues, while CKs and ABA did not. LM-array analysis revealed that sets of genes required for IAA and GA synthesis were coordinately expressed during the later stages of MS/POL development, suggesting that these genes are responsible for the massive accumulation of IAA and GA(4) in the mature anther. In contrast, genes for GA signaling were preferentially expressed during the early developmental stages of MS/POL and throughout TAP development, while their expression was down-regulated at the later stages of MS/POL development. In the case of auxin signaling genes, such mirror-imaged expression observed in GA synthesis and signaling genes was not observed. IAA receptor genes were mostly expressed during the late stages of MS/POL development, and various sets of AUX/IAA and ARF genes were expressed during the different stages of MS/POL or TAP development. Such cell type-specific expression profiles of phytohormone biosynthesis and signaling genes demonstrate the validity and importance of analyzing the expression of phytohormone-related genes in individual cell types independently of other cells/tissues.

Fig. 1

Representative gene expression profiles of the GA biosynthesis, deactivation and signaling pathways in the MS/POL and TAP of rice. Red and green colors indicate higher and lower expression, respectively. The color scale (representing the average log₂ Z-value normalized by R software) is shown at the bottom. The letters ‘Os’ at the beginning of each gene are omitted for convenience. MS/POL, microspore/pollen; TAP, tapetum; MEI, meiosis, TET, tetrad stage; UN, uninuclear MS stage; BC, bicellular POL stage; TC, tricellular POL stage.

Fig. 2

Cluster analysis of expression profiles (presented as log₂ Z-values) of rice GA biosynthesis, deactivation and signaling genes in the MS/POL and TAP. Genes were classified into Clusters I and II based on their specific expression patterns. Red and green colors indicate higher and lower expression, respectively. The color scale is shown at the bottom and the RAP-DB accession number and gene product names are shown on the right. EBC, early bicellular POL stage; MBC, middle bicellular POL stage; LBC, late bicellular POL stage. Other details are the same as in Fig. 1.

Fig. 3

Representative gene expression profiles of the IAA biosynthesis and signaling pathways in the MS/POL and TAP of rice. Other details are as in Fig. 1.

Fig. 4

Cluster analysis of expression profiles (presented as log₂ Z-values) of rice IAA biosynthesis and auxin signaling genes in the MS/POL and TAP. Genes were classified into Clusters I–IV based on their specific expression patterns. Other details are as in Fig. 1.

Fig. 5

Cluster analysis of expression profiles of rice IAA/AUX and ARF genes in the MS/POL and TAP. IAA/AUX genes are marked in light blue and ARF genes in dark blue. Genes were classified into Clusters I-1, I-2 and II-1 to II-5 based on their specific expression patterns. Other details are as in Fig. 1.

Fig. 6

Representative gene expression profiles of the CK biosynthesis, deactivation and signaling pathways in the MS/POL and TAP of rice. Other details are as in Fig. 1.

Fig. 7

Representative gene expression profiles of the BR biosynthesis, deactivation and signaling pathways in the MS/POL and TAP of rice. Other details are as in Fig. 1.

Fig. 8

Representative gene expression profiles of the ethylene biosynthesis and signaling pathways in the MS/POL and TAP of rice. Other details are as in Fig. 1.

Fig. 9

Representative gene expression profiles of the ABA biosynthesis, deactivation and signaling pathways in the MS/POL and TAP of rice. Other details are as in Fig. 1.

Fig. 10

Representative gene expression profiles of the JA biosynthesis and signaling pathways in the MS/POL and TAP of rice. Other details are as in Fig. 1.

Fig. 11

Schematic representation of changes in phytohormone levels in the MS/POL during development, as predicted from the microarray data. vn; vegetative nucleus, gc; generative cell.