Limits...
Involvements of PCD and changes in gene expression profile during self-pruning of spring shoots in sweet orange (Citrus sinensis).

Zhang JZ, Zhao K, Ai XY, Hu CG - BMC Genomics (2014)

Bottom Line: Our results indicated that PCD occurred systematically and progressively and may play an important role in the control of self-pruning of citrus.Some genes were related to PCD, while others were associated with cell wall biosynthesis or metabolism.Therefore, several key flowering homologs of Arabidopsis and tomato shoot apical meristem (SAM) activity genes were investigated in sweet orange by real-time PCR and in situ hybridization, and the results indicated that these genes were preferentially expressed in SAM as well as axillary meristem.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Horticultural Plant Biology (Ministry of Education), College of Horticulture and Forestry Science, Huazhong Agricultural University, Wuhan, 430070, China. chungen@mail.hzau.edu.cn.

ABSTRACT

Background: Citrus shoot tips abscise at an anatomically distinct abscission zone (AZ) that separates the top part of the shoots into basal and apical portions (citrus self-pruning). Cell separation occurs only at the AZ, which suggests its cells have distinctive molecular regulation. Although several studies have looked into the morphological aspects of self-pruning process, the underlying molecular mechanisms remain unknown.

Results: In this study, the hallmarks of programmed cell death (PCD) were identified by TUNEL experiments, transmission electron microscopy (TEM) and histochemical staining for reactive oxygen species (ROS) during self-pruning of the spring shoots in sweet orange. Our results indicated that PCD occurred systematically and progressively and may play an important role in the control of self-pruning of citrus. Microarray analysis was used to examine transcriptome changes at three stages of self-pruning, and 1,378 differentially expressed genes were identified. Some genes were related to PCD, while others were associated with cell wall biosynthesis or metabolism. These results strongly suggest that abscission layers activate both catabolic and anabolic wall modification pathways during the self-pruning process. In addition, a strong correlation was observed between self-pruning and the expression of hormone-related genes. Self-pruning plays an important role in citrus floral bud initiation. Therefore, several key flowering homologs of Arabidopsis and tomato shoot apical meristem (SAM) activity genes were investigated in sweet orange by real-time PCR and in situ hybridization, and the results indicated that these genes were preferentially expressed in SAM as well as axillary meristem.

Conclusion: Based on these findings, a model for sweet orange spring shoot self-pruning is proposed, which will enable us to better understand the mechanism of self-pruning and abscission.

Show MeSH
Cytological changes of ‘Cara Cara’ navel orange (Citrus sinensisOsbeck) spring shoot during self-pruning process by paraffin section and TEM analysis. Stages A–F occur before self-pruning, (A–F) 45 days, 35 days, 25 days, 15 days, 7 days, and 3 days before self-pruning of shoot tips, respectively. (G) 3 days before self-pruning of lateral bud; (H) 20 days after self-pruning; (I) lobular of shoot tips begin to fall (before self-pruning); (J) the appearance of the AZ; stages L–P occur after self-pruning, (K) visible AZ; (L, M) shoot tip begins to fall; (N) shoot tips after self-pruning; (O) before formation of protective layer of AZ; (P) after protective layer of AZ formed. Bars are 50 μm in L–P, and 100 μm in other photographs. Red arrows represent AZ.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4209071&req=5

Fig2: Cytological changes of ‘Cara Cara’ navel orange (Citrus sinensisOsbeck) spring shoot during self-pruning process by paraffin section and TEM analysis. Stages A–F occur before self-pruning, (A–F) 45 days, 35 days, 25 days, 15 days, 7 days, and 3 days before self-pruning of shoot tips, respectively. (G) 3 days before self-pruning of lateral bud; (H) 20 days after self-pruning; (I) lobular of shoot tips begin to fall (before self-pruning); (J) the appearance of the AZ; stages L–P occur after self-pruning, (K) visible AZ; (L, M) shoot tip begins to fall; (N) shoot tips after self-pruning; (O) before formation of protective layer of AZ; (P) after protective layer of AZ formed. Bars are 50 μm in L–P, and 100 μm in other photographs. Red arrows represent AZ.

Mentions: The paraffin sections and TEM analysis of shoot tip cells showed no visible evidence of cellular breakdown or death before self-pruning (Figures 2 and 3). Only after the shoot tip became yellow did the cytoplasm became less intensely stained compared to that in cells of the active growth regions (Figure 2A–E). This change in stainability may reflect an alteration in the chemical and/or physical nature of the protoplasm, and it was the first histological indication of an altered developmental pattern. Apices of the subjacent axillary buds would serve as the pseudoterminal bud (Figure 2G-H). In this bud, the cytoplasm became more densely stained (Figure 2H) than before self-pruning (Figure 2G). During the self-pruning process, when the shoots elongation was complete and lobular of shoot tips began to fall (Figure 2I), in the apical meristem of the unexpanded shoot tip cells became more vacuolated and their nuclei were condensed (Figures 2I and 3E). These changes suggested that a parallel senescence pattern was occurring, and scattered necrotic areas became evident in pith and cortex (Figure 2J), but at the abscission site, a separation layer in the stem was not yet apparent. After 2–3 days, the separation layer was visible (Figure 2K, L). At the later stage, when the tip including all the leaf primordia was completely necrotic, separation of the cells in pith and cortex of the stem at the abscission site had occurred (Figure 2M). The abscission site was commonly located in an internode distal to the sixth, seventh, or eighth leaf. No protective layer had formed in the stem at this stage (Figure 2N, O). After the shoot tip dropped off, the protective layer developed (Figure 2P).Figure 2


Involvements of PCD and changes in gene expression profile during self-pruning of spring shoots in sweet orange (Citrus sinensis).

Zhang JZ, Zhao K, Ai XY, Hu CG - BMC Genomics (2014)

Cytological changes of ‘Cara Cara’ navel orange (Citrus sinensisOsbeck) spring shoot during self-pruning process by paraffin section and TEM analysis. Stages A–F occur before self-pruning, (A–F) 45 days, 35 days, 25 days, 15 days, 7 days, and 3 days before self-pruning of shoot tips, respectively. (G) 3 days before self-pruning of lateral bud; (H) 20 days after self-pruning; (I) lobular of shoot tips begin to fall (before self-pruning); (J) the appearance of the AZ; stages L–P occur after self-pruning, (K) visible AZ; (L, M) shoot tip begins to fall; (N) shoot tips after self-pruning; (O) before formation of protective layer of AZ; (P) after protective layer of AZ formed. Bars are 50 μm in L–P, and 100 μm in other photographs. Red arrows represent AZ.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4209071&req=5

Fig2: Cytological changes of ‘Cara Cara’ navel orange (Citrus sinensisOsbeck) spring shoot during self-pruning process by paraffin section and TEM analysis. Stages A–F occur before self-pruning, (A–F) 45 days, 35 days, 25 days, 15 days, 7 days, and 3 days before self-pruning of shoot tips, respectively. (G) 3 days before self-pruning of lateral bud; (H) 20 days after self-pruning; (I) lobular of shoot tips begin to fall (before self-pruning); (J) the appearance of the AZ; stages L–P occur after self-pruning, (K) visible AZ; (L, M) shoot tip begins to fall; (N) shoot tips after self-pruning; (O) before formation of protective layer of AZ; (P) after protective layer of AZ formed. Bars are 50 μm in L–P, and 100 μm in other photographs. Red arrows represent AZ.
Mentions: The paraffin sections and TEM analysis of shoot tip cells showed no visible evidence of cellular breakdown or death before self-pruning (Figures 2 and 3). Only after the shoot tip became yellow did the cytoplasm became less intensely stained compared to that in cells of the active growth regions (Figure 2A–E). This change in stainability may reflect an alteration in the chemical and/or physical nature of the protoplasm, and it was the first histological indication of an altered developmental pattern. Apices of the subjacent axillary buds would serve as the pseudoterminal bud (Figure 2G-H). In this bud, the cytoplasm became more densely stained (Figure 2H) than before self-pruning (Figure 2G). During the self-pruning process, when the shoots elongation was complete and lobular of shoot tips began to fall (Figure 2I), in the apical meristem of the unexpanded shoot tip cells became more vacuolated and their nuclei were condensed (Figures 2I and 3E). These changes suggested that a parallel senescence pattern was occurring, and scattered necrotic areas became evident in pith and cortex (Figure 2J), but at the abscission site, a separation layer in the stem was not yet apparent. After 2–3 days, the separation layer was visible (Figure 2K, L). At the later stage, when the tip including all the leaf primordia was completely necrotic, separation of the cells in pith and cortex of the stem at the abscission site had occurred (Figure 2M). The abscission site was commonly located in an internode distal to the sixth, seventh, or eighth leaf. No protective layer had formed in the stem at this stage (Figure 2N, O). After the shoot tip dropped off, the protective layer developed (Figure 2P).Figure 2

Bottom Line: Our results indicated that PCD occurred systematically and progressively and may play an important role in the control of self-pruning of citrus.Some genes were related to PCD, while others were associated with cell wall biosynthesis or metabolism.Therefore, several key flowering homologs of Arabidopsis and tomato shoot apical meristem (SAM) activity genes were investigated in sweet orange by real-time PCR and in situ hybridization, and the results indicated that these genes were preferentially expressed in SAM as well as axillary meristem.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Horticultural Plant Biology (Ministry of Education), College of Horticulture and Forestry Science, Huazhong Agricultural University, Wuhan, 430070, China. chungen@mail.hzau.edu.cn.

ABSTRACT

Background: Citrus shoot tips abscise at an anatomically distinct abscission zone (AZ) that separates the top part of the shoots into basal and apical portions (citrus self-pruning). Cell separation occurs only at the AZ, which suggests its cells have distinctive molecular regulation. Although several studies have looked into the morphological aspects of self-pruning process, the underlying molecular mechanisms remain unknown.

Results: In this study, the hallmarks of programmed cell death (PCD) were identified by TUNEL experiments, transmission electron microscopy (TEM) and histochemical staining for reactive oxygen species (ROS) during self-pruning of the spring shoots in sweet orange. Our results indicated that PCD occurred systematically and progressively and may play an important role in the control of self-pruning of citrus. Microarray analysis was used to examine transcriptome changes at three stages of self-pruning, and 1,378 differentially expressed genes were identified. Some genes were related to PCD, while others were associated with cell wall biosynthesis or metabolism. These results strongly suggest that abscission layers activate both catabolic and anabolic wall modification pathways during the self-pruning process. In addition, a strong correlation was observed between self-pruning and the expression of hormone-related genes. Self-pruning plays an important role in citrus floral bud initiation. Therefore, several key flowering homologs of Arabidopsis and tomato shoot apical meristem (SAM) activity genes were investigated in sweet orange by real-time PCR and in situ hybridization, and the results indicated that these genes were preferentially expressed in SAM as well as axillary meristem.

Conclusion: Based on these findings, a model for sweet orange spring shoot self-pruning is proposed, which will enable us to better understand the mechanism of self-pruning and abscission.

Show MeSH