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Comparative transcriptome analysis reveals carbohydrate and lipid metabolism blocks in Brassica napus L. male sterility induced by the chemical hybridization agent monosulfuron ester sodium.

Li Z, Cheng Y, Cui J, Zhang P, Zhao H, Hu S - BMC Genomics (2015)

Bottom Line: Transcripts involved in metabolism, especially in carbohydrate and lipid metabolism, and cellular transport were differentially expressed.Pathway visualization showed that the tightly regulated gene network for metabolism was reprogrammed to respond to MES treatment.MES treatment led to decrease in soluble sugars content in leaves and early stage buds, but increase in soluble sugars content and decrease in starch content in middle stage buds.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Crop Stress Biology in Arid Areas, Northwest A&F University, Yangling, Shaanxi, 712100, P. R. China. lzj8705@163.com.

ABSTRACT

Background: Chemical hybridization agents (CHAs) are often used to induce male sterility for the production of hybrid seeds. We previously discovered that monosulfuron ester sodium (MES), an acetolactate synthase (ALS) inhibitor of the herbicide sulfonylurea family, can induce rapeseed (Brassica napus L.) male sterility at approximately 1% concentration required for its herbicidal activity. To find some clues to the mechanism of MES inducing male sterility, the ultrastructural cytology observations, comparative transcriptome analysis, and physiological analysis on carbohydrate content were carried out in leaves and anthers at different developmental stages between the MES-treated and mock-treated rapeseed plants.

Results: Cytological analysis revealed that the plastid ultrastructure was abnormal in pollen mother cells and tapetal cells in male sterility anthers induced by MES treatment, with less material accumulation in it. However, starch granules were observed in chloroplastids of the epidermis cells in male sterility anthers. Comparative transcriptome analysis identified 1501 differentially expressed transcripts (DETs) in leaves and anthers at different developmental stages, most of these DETs being localized in plastid and mitochondrion. Transcripts involved in metabolism, especially in carbohydrate and lipid metabolism, and cellular transport were differentially expressed. Pathway visualization showed that the tightly regulated gene network for metabolism was reprogrammed to respond to MES treatment. The results of cytological observation and transcriptome analysis in the MES-treated rapeseed plants were mirrored by carbohydrate content analysis. MES treatment led to decrease in soluble sugars content in leaves and early stage buds, but increase in soluble sugars content and decrease in starch content in middle stage buds.

Conclusions: Our integrative results suggested that carbohydrate and lipid metabolism were influenced by CHA-MES treatment during rapeseed anther development, which might responsible for low concentration MES specifically inducing male sterility. A simple action model of CHA-MES inducing male sterility in B. napus was proposed. These results will help us to understand the mechanism of MES inducing male sterility at low concentration, and might provide some potential targets for developing new male sterility inducing CHAs and for genetic manipulation in rapeseed breeding.

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Related in: MedlinePlus

Transmission Electron Microscope (TEM) micrographs of the anthers from the mock-treated (fertile) and MES-treated (sterile) plants. (A) The fertile anthers at pollen mother cell (PMC) stage; (B) Enlarged fertile meiocytes in (A); and (C) Enlarged fertile tapetum in (A) showing numerous plastids dispersed in cytoplasm (white arrow). (D) The sterile anthers at PMC stage; (E) Enlarged sterile meiocytes in (D) showing less plastids in condensed cytoplasm separated from the cell wall; (F) Enlarged sterile tapetum in (D) showing little abnormal plastids (white arrow) and more large vacuoles in cytoplasm, and with a little plasmolysis at meiocyte side (black arrow). (G) The fertile anthers at vacuolated-microspore stage; (H) The degraded tapetum in (G) showing elaioplasts and tapetsomes with abundant lipids; (I) Plastids in tapetum located in a crown showing filled with globular low electron-dense metabolites and surrounded by rich endoplasmic reticulum (ER). (J) The sterile anthers at vacuolated-microspore stage (type I); (K) The undegraded tapetum in (J) showing elaioplasts and tapetsomes with abundant lipids; (L) Plastids in tapetum located in a crown showing irregular shaped low electron-dense material. (M) The sterile anthers at vacuolated-microspore stage (type II); (N) The degraded tapetum in (M) showing scattered elaioplasts and tapetsomes with fuzzy structure; (O) The fertile anthers at mature pollen grain stage; (P) The pollen grain in (O) showing profuse globular particles; (Q) The enlarged globular particles in (P). (R) The sterile anthers at mature pollen grain stage (type II); (S) The undegraded tapetum in (R) died but cell wall still existed (black arrow); (T) The sterile anthers at mature pollen grain stage (type I). (U) The epidermis and endothecium cells in fertile plants at vacuolated-microspore stage; (V) The epidermis cells in (U) showing normal oval-shaped chloroplastids with distinct thylakoid structure and little starch granules in thylakoid; (W) The endothecium cells in (U) showing oval-shaped chloroplastids with distinct thylakoid structure. (X) The epidermis and endothecium cells in sterile plants at vacuolated-microspore stage; (Y) The epidermis cells in (X) showing abnormal chloroplastids with large starch granules in thylakoid; (Z) The endothecium cells in (X) showing fusiform-shaped chloroplastids with linear thylakoid structure. PMC, pollen mother cell; N, nucleus; T, tapetum; Msp, microspore; Ep, elaioplast; Ts, tapetosome; PG, pollen grain; TCW, tapetum cell wall; E, epidermis; En, endothecium; Ch, chloroplast. Scale bars = 10 μm (A, D, G, J, M, N, O and T), 5 μm (C, F, P, R, S, U and X), 2 μm (B, E and K), and 1 μm (H, I, L, Q, V, W, Y and Z).
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Fig1: Transmission Electron Microscope (TEM) micrographs of the anthers from the mock-treated (fertile) and MES-treated (sterile) plants. (A) The fertile anthers at pollen mother cell (PMC) stage; (B) Enlarged fertile meiocytes in (A); and (C) Enlarged fertile tapetum in (A) showing numerous plastids dispersed in cytoplasm (white arrow). (D) The sterile anthers at PMC stage; (E) Enlarged sterile meiocytes in (D) showing less plastids in condensed cytoplasm separated from the cell wall; (F) Enlarged sterile tapetum in (D) showing little abnormal plastids (white arrow) and more large vacuoles in cytoplasm, and with a little plasmolysis at meiocyte side (black arrow). (G) The fertile anthers at vacuolated-microspore stage; (H) The degraded tapetum in (G) showing elaioplasts and tapetsomes with abundant lipids; (I) Plastids in tapetum located in a crown showing filled with globular low electron-dense metabolites and surrounded by rich endoplasmic reticulum (ER). (J) The sterile anthers at vacuolated-microspore stage (type I); (K) The undegraded tapetum in (J) showing elaioplasts and tapetsomes with abundant lipids; (L) Plastids in tapetum located in a crown showing irregular shaped low electron-dense material. (M) The sterile anthers at vacuolated-microspore stage (type II); (N) The degraded tapetum in (M) showing scattered elaioplasts and tapetsomes with fuzzy structure; (O) The fertile anthers at mature pollen grain stage; (P) The pollen grain in (O) showing profuse globular particles; (Q) The enlarged globular particles in (P). (R) The sterile anthers at mature pollen grain stage (type II); (S) The undegraded tapetum in (R) died but cell wall still existed (black arrow); (T) The sterile anthers at mature pollen grain stage (type I). (U) The epidermis and endothecium cells in fertile plants at vacuolated-microspore stage; (V) The epidermis cells in (U) showing normal oval-shaped chloroplastids with distinct thylakoid structure and little starch granules in thylakoid; (W) The endothecium cells in (U) showing oval-shaped chloroplastids with distinct thylakoid structure. (X) The epidermis and endothecium cells in sterile plants at vacuolated-microspore stage; (Y) The epidermis cells in (X) showing abnormal chloroplastids with large starch granules in thylakoid; (Z) The endothecium cells in (X) showing fusiform-shaped chloroplastids with linear thylakoid structure. PMC, pollen mother cell; N, nucleus; T, tapetum; Msp, microspore; Ep, elaioplast; Ts, tapetosome; PG, pollen grain; TCW, tapetum cell wall; E, epidermis; En, endothecium; Ch, chloroplast. Scale bars = 10 μm (A, D, G, J, M, N, O and T), 5 μm (C, F, P, R, S, U and X), 2 μm (B, E and K), and 1 μm (H, I, L, Q, V, W, Y and Z).

Mentions: We previously showed that MES treatment causes two typical defects in sterile anthers: type I with early broken down tapetum at the PMC stage and type II with abnormal nondegraded tapetum at the mature pollen stage [8]. To better understand these phenomena, we observed the ultrastructure of fertile and sterile anthers from the mock-treated and MES-treated plants, respectively, during their development. The results showed that MES treatment affected the plastid ultrastructure and metabolite accumulation in the developing anthers (Figure 1). At the PMC stage, numerous plastids are dispersed in the cytoplasm of PMCs and tapetal cells in the mock-treated plants (Figure 1A–C). However, serious plasmolysis in PMCs and slight plasmolysis in tapetal cells were observed in the MES-treated male sterile plants (Figure 1D, E, and black arrow in 1F), and the cytoplasm of meiocytes and tapetal cells exhibited low electron density, with less plastids dispersed in them. At the vacuolated-microspore stage, the tapetum cells began to degrade and a number of elaioplasts and tapetosomes with abundant lipid compounds were formed in the tapetum of the mock-treated fertile anther (Figure 1G, H, white arrow). Besides, another type of plastids located in a crown, started to accumulate low electron-dense material and was surrounded by the rich ER (Figure 1I, white arrow). In contrast, in the MES-treated sterile plants, two types of abnormal tapetum were observed, as shown by Cheng et al. (2013) [8]. In type II abnormal tapetum, a number of elaioplasts and tapetosomes were formed, as seen in fertile plants; however, the tapetum cells did not degrade, the crowned plastids showed an irregular shape and were not well developed (Figure 1J, K, L). Type I abnormal tapetum was degraded and released noncompact elaioplasts and low electron-dense tapetosomes (Figure 1M, N). At the mature-pollen stage, the fertile pollen grains showed profuse globular particles (Figure 1O, P, Q); however, the sterile pollen grains were almost empty, type II tapetum still showed an intact and visible tapetal cell wall (Figure 1R, S), and type I tapetum showed solidified bulks (Figure 1T).Figure 1


Comparative transcriptome analysis reveals carbohydrate and lipid metabolism blocks in Brassica napus L. male sterility induced by the chemical hybridization agent monosulfuron ester sodium.

Li Z, Cheng Y, Cui J, Zhang P, Zhao H, Hu S - BMC Genomics (2015)

Transmission Electron Microscope (TEM) micrographs of the anthers from the mock-treated (fertile) and MES-treated (sterile) plants. (A) The fertile anthers at pollen mother cell (PMC) stage; (B) Enlarged fertile meiocytes in (A); and (C) Enlarged fertile tapetum in (A) showing numerous plastids dispersed in cytoplasm (white arrow). (D) The sterile anthers at PMC stage; (E) Enlarged sterile meiocytes in (D) showing less plastids in condensed cytoplasm separated from the cell wall; (F) Enlarged sterile tapetum in (D) showing little abnormal plastids (white arrow) and more large vacuoles in cytoplasm, and with a little plasmolysis at meiocyte side (black arrow). (G) The fertile anthers at vacuolated-microspore stage; (H) The degraded tapetum in (G) showing elaioplasts and tapetsomes with abundant lipids; (I) Plastids in tapetum located in a crown showing filled with globular low electron-dense metabolites and surrounded by rich endoplasmic reticulum (ER). (J) The sterile anthers at vacuolated-microspore stage (type I); (K) The undegraded tapetum in (J) showing elaioplasts and tapetsomes with abundant lipids; (L) Plastids in tapetum located in a crown showing irregular shaped low electron-dense material. (M) The sterile anthers at vacuolated-microspore stage (type II); (N) The degraded tapetum in (M) showing scattered elaioplasts and tapetsomes with fuzzy structure; (O) The fertile anthers at mature pollen grain stage; (P) The pollen grain in (O) showing profuse globular particles; (Q) The enlarged globular particles in (P). (R) The sterile anthers at mature pollen grain stage (type II); (S) The undegraded tapetum in (R) died but cell wall still existed (black arrow); (T) The sterile anthers at mature pollen grain stage (type I). (U) The epidermis and endothecium cells in fertile plants at vacuolated-microspore stage; (V) The epidermis cells in (U) showing normal oval-shaped chloroplastids with distinct thylakoid structure and little starch granules in thylakoid; (W) The endothecium cells in (U) showing oval-shaped chloroplastids with distinct thylakoid structure. (X) The epidermis and endothecium cells in sterile plants at vacuolated-microspore stage; (Y) The epidermis cells in (X) showing abnormal chloroplastids with large starch granules in thylakoid; (Z) The endothecium cells in (X) showing fusiform-shaped chloroplastids with linear thylakoid structure. PMC, pollen mother cell; N, nucleus; T, tapetum; Msp, microspore; Ep, elaioplast; Ts, tapetosome; PG, pollen grain; TCW, tapetum cell wall; E, epidermis; En, endothecium; Ch, chloroplast. Scale bars = 10 μm (A, D, G, J, M, N, O and T), 5 μm (C, F, P, R, S, U and X), 2 μm (B, E and K), and 1 μm (H, I, L, Q, V, W, Y and Z).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Fig1: Transmission Electron Microscope (TEM) micrographs of the anthers from the mock-treated (fertile) and MES-treated (sterile) plants. (A) The fertile anthers at pollen mother cell (PMC) stage; (B) Enlarged fertile meiocytes in (A); and (C) Enlarged fertile tapetum in (A) showing numerous plastids dispersed in cytoplasm (white arrow). (D) The sterile anthers at PMC stage; (E) Enlarged sterile meiocytes in (D) showing less plastids in condensed cytoplasm separated from the cell wall; (F) Enlarged sterile tapetum in (D) showing little abnormal plastids (white arrow) and more large vacuoles in cytoplasm, and with a little plasmolysis at meiocyte side (black arrow). (G) The fertile anthers at vacuolated-microspore stage; (H) The degraded tapetum in (G) showing elaioplasts and tapetsomes with abundant lipids; (I) Plastids in tapetum located in a crown showing filled with globular low electron-dense metabolites and surrounded by rich endoplasmic reticulum (ER). (J) The sterile anthers at vacuolated-microspore stage (type I); (K) The undegraded tapetum in (J) showing elaioplasts and tapetsomes with abundant lipids; (L) Plastids in tapetum located in a crown showing irregular shaped low electron-dense material. (M) The sterile anthers at vacuolated-microspore stage (type II); (N) The degraded tapetum in (M) showing scattered elaioplasts and tapetsomes with fuzzy structure; (O) The fertile anthers at mature pollen grain stage; (P) The pollen grain in (O) showing profuse globular particles; (Q) The enlarged globular particles in (P). (R) The sterile anthers at mature pollen grain stage (type II); (S) The undegraded tapetum in (R) died but cell wall still existed (black arrow); (T) The sterile anthers at mature pollen grain stage (type I). (U) The epidermis and endothecium cells in fertile plants at vacuolated-microspore stage; (V) The epidermis cells in (U) showing normal oval-shaped chloroplastids with distinct thylakoid structure and little starch granules in thylakoid; (W) The endothecium cells in (U) showing oval-shaped chloroplastids with distinct thylakoid structure. (X) The epidermis and endothecium cells in sterile plants at vacuolated-microspore stage; (Y) The epidermis cells in (X) showing abnormal chloroplastids with large starch granules in thylakoid; (Z) The endothecium cells in (X) showing fusiform-shaped chloroplastids with linear thylakoid structure. PMC, pollen mother cell; N, nucleus; T, tapetum; Msp, microspore; Ep, elaioplast; Ts, tapetosome; PG, pollen grain; TCW, tapetum cell wall; E, epidermis; En, endothecium; Ch, chloroplast. Scale bars = 10 μm (A, D, G, J, M, N, O and T), 5 μm (C, F, P, R, S, U and X), 2 μm (B, E and K), and 1 μm (H, I, L, Q, V, W, Y and Z).
Mentions: We previously showed that MES treatment causes two typical defects in sterile anthers: type I with early broken down tapetum at the PMC stage and type II with abnormal nondegraded tapetum at the mature pollen stage [8]. To better understand these phenomena, we observed the ultrastructure of fertile and sterile anthers from the mock-treated and MES-treated plants, respectively, during their development. The results showed that MES treatment affected the plastid ultrastructure and metabolite accumulation in the developing anthers (Figure 1). At the PMC stage, numerous plastids are dispersed in the cytoplasm of PMCs and tapetal cells in the mock-treated plants (Figure 1A–C). However, serious plasmolysis in PMCs and slight plasmolysis in tapetal cells were observed in the MES-treated male sterile plants (Figure 1D, E, and black arrow in 1F), and the cytoplasm of meiocytes and tapetal cells exhibited low electron density, with less plastids dispersed in them. At the vacuolated-microspore stage, the tapetum cells began to degrade and a number of elaioplasts and tapetosomes with abundant lipid compounds were formed in the tapetum of the mock-treated fertile anther (Figure 1G, H, white arrow). Besides, another type of plastids located in a crown, started to accumulate low electron-dense material and was surrounded by the rich ER (Figure 1I, white arrow). In contrast, in the MES-treated sterile plants, two types of abnormal tapetum were observed, as shown by Cheng et al. (2013) [8]. In type II abnormal tapetum, a number of elaioplasts and tapetosomes were formed, as seen in fertile plants; however, the tapetum cells did not degrade, the crowned plastids showed an irregular shape and were not well developed (Figure 1J, K, L). Type I abnormal tapetum was degraded and released noncompact elaioplasts and low electron-dense tapetosomes (Figure 1M, N). At the mature-pollen stage, the fertile pollen grains showed profuse globular particles (Figure 1O, P, Q); however, the sterile pollen grains were almost empty, type II tapetum still showed an intact and visible tapetal cell wall (Figure 1R, S), and type I tapetum showed solidified bulks (Figure 1T).Figure 1

Bottom Line: Transcripts involved in metabolism, especially in carbohydrate and lipid metabolism, and cellular transport were differentially expressed.Pathway visualization showed that the tightly regulated gene network for metabolism was reprogrammed to respond to MES treatment.MES treatment led to decrease in soluble sugars content in leaves and early stage buds, but increase in soluble sugars content and decrease in starch content in middle stage buds.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Crop Stress Biology in Arid Areas, Northwest A&F University, Yangling, Shaanxi, 712100, P. R. China. lzj8705@163.com.

ABSTRACT

Background: Chemical hybridization agents (CHAs) are often used to induce male sterility for the production of hybrid seeds. We previously discovered that monosulfuron ester sodium (MES), an acetolactate synthase (ALS) inhibitor of the herbicide sulfonylurea family, can induce rapeseed (Brassica napus L.) male sterility at approximately 1% concentration required for its herbicidal activity. To find some clues to the mechanism of MES inducing male sterility, the ultrastructural cytology observations, comparative transcriptome analysis, and physiological analysis on carbohydrate content were carried out in leaves and anthers at different developmental stages between the MES-treated and mock-treated rapeseed plants.

Results: Cytological analysis revealed that the plastid ultrastructure was abnormal in pollen mother cells and tapetal cells in male sterility anthers induced by MES treatment, with less material accumulation in it. However, starch granules were observed in chloroplastids of the epidermis cells in male sterility anthers. Comparative transcriptome analysis identified 1501 differentially expressed transcripts (DETs) in leaves and anthers at different developmental stages, most of these DETs being localized in plastid and mitochondrion. Transcripts involved in metabolism, especially in carbohydrate and lipid metabolism, and cellular transport were differentially expressed. Pathway visualization showed that the tightly regulated gene network for metabolism was reprogrammed to respond to MES treatment. The results of cytological observation and transcriptome analysis in the MES-treated rapeseed plants were mirrored by carbohydrate content analysis. MES treatment led to decrease in soluble sugars content in leaves and early stage buds, but increase in soluble sugars content and decrease in starch content in middle stage buds.

Conclusions: Our integrative results suggested that carbohydrate and lipid metabolism were influenced by CHA-MES treatment during rapeseed anther development, which might responsible for low concentration MES specifically inducing male sterility. A simple action model of CHA-MES inducing male sterility in B. napus was proposed. These results will help us to understand the mechanism of MES inducing male sterility at low concentration, and might provide some potential targets for developing new male sterility inducing CHAs and for genetic manipulation in rapeseed breeding.

Show MeSH
Related in: MedlinePlus