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OsABCG15 encodes a membrane protein that plays an important role in anther cuticle and pollen exine formation in rice.

Wu L, Guan Y, Wu Z, Yang K, Lv J, Converse R, Huang Y, Mao J, Zhao Y, Wang Z, Min H, Kan D, Zhang Y - Plant Cell Rep. (2014)

Bottom Line: Using map-based cloning, we found a spontaneous A-to-C transition in the fourth exon of OsABCG15 that caused an amino acid substitution of Thr-to-Pro in the predicted ATP-binding domain of the protein sequence.Our results suggested that OsABCG15 played an essential role in the formation of the rice anther cuticle and pollen exine.This role may include the secretion of the lipid precursors from the tapetum to facilitate the transfer of precursors to the surface of the anther epidermis as well as to microspores.

View Article: PubMed Central - PubMed

Affiliation: College of Agronomy and Biotechnology, Southwest University, Chongqing, 400715, China.

ABSTRACT

Key message: An ABC transporter gene ( OsABCG15 ) was proven to be involved in pollen development in rice. The corresponding protein was localized on the plasma membrane using subcellular localization. Wax, cutin, and sporopollenin are important for normal development of the anther cuticle and pollen exine, respectively. Their lipid soluble precursors, which are produced in the tapetum, are then secreted and transferred to the anther and microspore surface for polymerization. However, little is known about the mechanisms underlying the transport of these precursors. Here, we identified and characterized a member of the G subfamily of ATP-binding cassette (ABC) transporters, OsABCG15, which is required for the secretion of these lipid-soluble precursors in rice. Using map-based cloning, we found a spontaneous A-to-C transition in the fourth exon of OsABCG15 that caused an amino acid substitution of Thr-to-Pro in the predicted ATP-binding domain of the protein sequence. This osabcg15 mutant failed to produce any viable pollen and was completely male sterile. Histological analysis indicated that osabcg15 exhibited an undeveloped anther cuticle, enlarged middle layer, abnormal Ubisch body development, tapetum degeneration with a falling apart style, and collapsed pollen grains without detectable exine. OsABCG15 was expressed preferentially in the tapetum, and the fused GFP-OsABCG15 protein was localized to the plasma membrane. Our results suggested that OsABCG15 played an essential role in the formation of the rice anther cuticle and pollen exine. This role may include the secretion of the lipid precursors from the tapetum to facilitate the transfer of precursors to the surface of the anther epidermis as well as to microspores.

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Comparison of anther and pollen between the wild type and osabcg15 by transmission electron microscopy. Ba bacula, C cuticle, CW cell wall, DMsp degenerated microspore, DPE degenerated prim-exine, EX exine, Msp microspore, Ne nexine, PMC pollen mother cell, PE prim-exine, Te tectum, Tds tetrads, TMR tapetum and microspore residue, TW tapetum cell wall, Ub Ubisch, V vacuole. Bars 1 μm in A–L, and a–l; 200 nm in M–R, and m–r. Cross-sections of the wild-type tapetum at stages 7 (A), 8a (B), 8b (C), 9 (D), 10 (E), and 11 (F). Cross-sections of the osabcg15 tapetum at stages 7 (a), 8a (b), 8b (c), 9 (d), 10 (e) and 11 (f). G–L The pollen exine development of the wild type from stages 7–11. g–l Defective pollen exine development of osabcg15 from stages 7–11. M–R Outer region of anther epidermis in the wild type from stages 7–11. m–r Outer region of the anther epidermis in the osabcg15 mutant from stages 7–11
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Fig4: Comparison of anther and pollen between the wild type and osabcg15 by transmission electron microscopy. Ba bacula, C cuticle, CW cell wall, DMsp degenerated microspore, DPE degenerated prim-exine, EX exine, Msp microspore, Ne nexine, PMC pollen mother cell, PE prim-exine, Te tectum, Tds tetrads, TMR tapetum and microspore residue, TW tapetum cell wall, Ub Ubisch, V vacuole. Bars 1 μm in A–L, and a–l; 200 nm in M–R, and m–r. Cross-sections of the wild-type tapetum at stages 7 (A), 8a (B), 8b (C), 9 (D), 10 (E), and 11 (F). Cross-sections of the osabcg15 tapetum at stages 7 (a), 8a (b), 8b (c), 9 (d), 10 (e) and 11 (f). G–L The pollen exine development of the wild type from stages 7–11. g–l Defective pollen exine development of osabcg15 from stages 7–11. M–R Outer region of anther epidermis in the wild type from stages 7–11. m–r Outer region of the anther epidermis in the osabcg15 mutant from stages 7–11

Mentions: To improve our understanding about the abnormalities of the internal anther wall and pollen cells in osabcg15, TEM was performed. Using the same procedure as previously described using semi-sections of anther, we confirmed the abnormal widening of the middle layer and incomplete degeneration of the endothecium in osabcg15 (see Supplementary Fig. 1 online). The tapetum and microspores were then compared. In agreement with the semi-section and SEM results, during stages 7–8b, the microspores in osabcg15 were more frequently wizened than those in the wild type (see Supplementary Fig. 2 online). No significant differences were detected in the tapetum and microspore structures when comparing the wild-type and osabcg15 (Fig. 4A–C, G–I, a–c, g–i). At stage 9, numerous prim-Ubisch, intermediate to small chifres, were released onto the peripheral side of the tapetum in wild type, indicating the initiation of the secretion of sporopollenin precursor from the sporophytic tapetum (Fig. 4D). Meanwhile, the wild-type microspores formed a primary exine structure, composed of the nexine and baculum, and had a round shape (Fig. 4J). In contrast, the osabcg15 Ubisch bodies were smaller and failed to break through the tapetal cell wall even though they were created in stage 8b (Fig. 4c, d). At the same time, the exine of osabcg15 microspores failed to initiate deposition of the sporopollenin-composing materials, and the surfaces of some microspores looked wavy (Fig. 4j; see Supplementary Fig. 2 online). At stage 10 in wild type, the tapetum became condensed along with high vacuolization and the continuation of degradation, while the Ubisch bodies expanded (Fig. 4E). Because of the increased amount of precursors depositing onto the microspore surfaces and then polymerizing, the microspores formed a regular exine structure with nexine, baculum, and tectum on the surface (Fig. 4K). However, the whole tapetum of osabcg15, including the peripheral region, dispersed completely, and the immature Ubisch bodies beneath the tapetum cell wall were destroyed (Fig. 4e), indicating serious defects in the synthesis and transport of sporopollenin precursors. Instead of developing into the classical exine bilayer, the thin primexine degraded from a lack of sporopollenin components, and the microspores eventually collapsed (Fig. 4k). In the final stages, except for stage 11, degeneration continued, and the tapetum of wild type was left with just a thin layer of peripheral region to hold the mature Ubisch bodies (Fig. 4F). The microspore exine thickened from the abundant deposition of sporopollenin (Fig. 4L). However, during these stages in osabcg15, there was total degeneration of the tapetum and microspores, and no clear structures of tapetum, Ubisch bodies, or microspores could be found in the shriveled locules except for residues produced by the degraded tapetum and microspores (Fig. 4f, l).Fig. 4


OsABCG15 encodes a membrane protein that plays an important role in anther cuticle and pollen exine formation in rice.

Wu L, Guan Y, Wu Z, Yang K, Lv J, Converse R, Huang Y, Mao J, Zhao Y, Wang Z, Min H, Kan D, Zhang Y - Plant Cell Rep. (2014)

Comparison of anther and pollen between the wild type and osabcg15 by transmission electron microscopy. Ba bacula, C cuticle, CW cell wall, DMsp degenerated microspore, DPE degenerated prim-exine, EX exine, Msp microspore, Ne nexine, PMC pollen mother cell, PE prim-exine, Te tectum, Tds tetrads, TMR tapetum and microspore residue, TW tapetum cell wall, Ub Ubisch, V vacuole. Bars 1 μm in A–L, and a–l; 200 nm in M–R, and m–r. Cross-sections of the wild-type tapetum at stages 7 (A), 8a (B), 8b (C), 9 (D), 10 (E), and 11 (F). Cross-sections of the osabcg15 tapetum at stages 7 (a), 8a (b), 8b (c), 9 (d), 10 (e) and 11 (f). G–L The pollen exine development of the wild type from stages 7–11. g–l Defective pollen exine development of osabcg15 from stages 7–11. M–R Outer region of anther epidermis in the wild type from stages 7–11. m–r Outer region of the anther epidermis in the osabcg15 mutant from stages 7–11
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Fig4: Comparison of anther and pollen between the wild type and osabcg15 by transmission electron microscopy. Ba bacula, C cuticle, CW cell wall, DMsp degenerated microspore, DPE degenerated prim-exine, EX exine, Msp microspore, Ne nexine, PMC pollen mother cell, PE prim-exine, Te tectum, Tds tetrads, TMR tapetum and microspore residue, TW tapetum cell wall, Ub Ubisch, V vacuole. Bars 1 μm in A–L, and a–l; 200 nm in M–R, and m–r. Cross-sections of the wild-type tapetum at stages 7 (A), 8a (B), 8b (C), 9 (D), 10 (E), and 11 (F). Cross-sections of the osabcg15 tapetum at stages 7 (a), 8a (b), 8b (c), 9 (d), 10 (e) and 11 (f). G–L The pollen exine development of the wild type from stages 7–11. g–l Defective pollen exine development of osabcg15 from stages 7–11. M–R Outer region of anther epidermis in the wild type from stages 7–11. m–r Outer region of the anther epidermis in the osabcg15 mutant from stages 7–11
Mentions: To improve our understanding about the abnormalities of the internal anther wall and pollen cells in osabcg15, TEM was performed. Using the same procedure as previously described using semi-sections of anther, we confirmed the abnormal widening of the middle layer and incomplete degeneration of the endothecium in osabcg15 (see Supplementary Fig. 1 online). The tapetum and microspores were then compared. In agreement with the semi-section and SEM results, during stages 7–8b, the microspores in osabcg15 were more frequently wizened than those in the wild type (see Supplementary Fig. 2 online). No significant differences were detected in the tapetum and microspore structures when comparing the wild-type and osabcg15 (Fig. 4A–C, G–I, a–c, g–i). At stage 9, numerous prim-Ubisch, intermediate to small chifres, were released onto the peripheral side of the tapetum in wild type, indicating the initiation of the secretion of sporopollenin precursor from the sporophytic tapetum (Fig. 4D). Meanwhile, the wild-type microspores formed a primary exine structure, composed of the nexine and baculum, and had a round shape (Fig. 4J). In contrast, the osabcg15 Ubisch bodies were smaller and failed to break through the tapetal cell wall even though they were created in stage 8b (Fig. 4c, d). At the same time, the exine of osabcg15 microspores failed to initiate deposition of the sporopollenin-composing materials, and the surfaces of some microspores looked wavy (Fig. 4j; see Supplementary Fig. 2 online). At stage 10 in wild type, the tapetum became condensed along with high vacuolization and the continuation of degradation, while the Ubisch bodies expanded (Fig. 4E). Because of the increased amount of precursors depositing onto the microspore surfaces and then polymerizing, the microspores formed a regular exine structure with nexine, baculum, and tectum on the surface (Fig. 4K). However, the whole tapetum of osabcg15, including the peripheral region, dispersed completely, and the immature Ubisch bodies beneath the tapetum cell wall were destroyed (Fig. 4e), indicating serious defects in the synthesis and transport of sporopollenin precursors. Instead of developing into the classical exine bilayer, the thin primexine degraded from a lack of sporopollenin components, and the microspores eventually collapsed (Fig. 4k). In the final stages, except for stage 11, degeneration continued, and the tapetum of wild type was left with just a thin layer of peripheral region to hold the mature Ubisch bodies (Fig. 4F). The microspore exine thickened from the abundant deposition of sporopollenin (Fig. 4L). However, during these stages in osabcg15, there was total degeneration of the tapetum and microspores, and no clear structures of tapetum, Ubisch bodies, or microspores could be found in the shriveled locules except for residues produced by the degraded tapetum and microspores (Fig. 4f, l).Fig. 4

Bottom Line: Using map-based cloning, we found a spontaneous A-to-C transition in the fourth exon of OsABCG15 that caused an amino acid substitution of Thr-to-Pro in the predicted ATP-binding domain of the protein sequence.Our results suggested that OsABCG15 played an essential role in the formation of the rice anther cuticle and pollen exine.This role may include the secretion of the lipid precursors from the tapetum to facilitate the transfer of precursors to the surface of the anther epidermis as well as to microspores.

View Article: PubMed Central - PubMed

Affiliation: College of Agronomy and Biotechnology, Southwest University, Chongqing, 400715, China.

ABSTRACT

Key message: An ABC transporter gene ( OsABCG15 ) was proven to be involved in pollen development in rice. The corresponding protein was localized on the plasma membrane using subcellular localization. Wax, cutin, and sporopollenin are important for normal development of the anther cuticle and pollen exine, respectively. Their lipid soluble precursors, which are produced in the tapetum, are then secreted and transferred to the anther and microspore surface for polymerization. However, little is known about the mechanisms underlying the transport of these precursors. Here, we identified and characterized a member of the G subfamily of ATP-binding cassette (ABC) transporters, OsABCG15, which is required for the secretion of these lipid-soluble precursors in rice. Using map-based cloning, we found a spontaneous A-to-C transition in the fourth exon of OsABCG15 that caused an amino acid substitution of Thr-to-Pro in the predicted ATP-binding domain of the protein sequence. This osabcg15 mutant failed to produce any viable pollen and was completely male sterile. Histological analysis indicated that osabcg15 exhibited an undeveloped anther cuticle, enlarged middle layer, abnormal Ubisch body development, tapetum degeneration with a falling apart style, and collapsed pollen grains without detectable exine. OsABCG15 was expressed preferentially in the tapetum, and the fused GFP-OsABCG15 protein was localized to the plasma membrane. Our results suggested that OsABCG15 played an essential role in the formation of the rice anther cuticle and pollen exine. This role may include the secretion of the lipid precursors from the tapetum to facilitate the transfer of precursors to the surface of the anther epidermis as well as to microspores.

Show MeSH
Related in: MedlinePlus