Limits...
Cotyledon cells of Vigna mungo seedlings use at least two distinct autophagic machineries for degradation of starch granules and cellular components.

Toyooka K, Okamoto T, Minamikawa T - J. Cell Biol. (2001)

Bottom Line: The results revealed that SG is inserted into LV through autophagic function of LV and subsequently degraded by vacuolar alpha-amylase.When the embryo axes were removed from seeds and the detached cotyledons were incubated, the autophagosome-mediated autophagy was observed, but the autophagic process for the degradation of SG was not detected, suggesting that these two autophagic processes were mediated by different cellular mechanisms.The two distinct autophagic processes were thought to be involved in the breakdown of SG and cell components in the cells of germinated cotyledon.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Tokyo Metropolitan University, Tokyo, 192-0397 Japan.

ABSTRACT
alpha-Amylase is expressed in cotyledons of germinated Vigna mungo seeds and is responsible for the degradation of starch that is stored in the starch granule (SG). Immunocytochemical analysis of the cotyledon cells with anti-alpha-amylase antibody showed that alpha-amylase is transported to protein storage vacuole (PSV) and lytic vacuole (LV), which is converted from PSV by hydrolysis of storage proteins. To observe the insertion/degradation processes of SG into/in the inside of vacuoles, ultrastructural analyses of the cotyledon cells were conducted. The results revealed that SG is inserted into LV through autophagic function of LV and subsequently degraded by vacuolar alpha-amylase. The autophagy for SG was structurally similar to micropexophagy detected in yeast cells. In addition to the autophagic process for SG, autophagosome-mediated autophagy for cytoplasm and mitochondria was detected in the cotyledon cells. When the embryo axes were removed from seeds and the detached cotyledons were incubated, the autophagosome-mediated autophagy was observed, but the autophagic process for the degradation of SG was not detected, suggesting that these two autophagic processes were mediated by different cellular mechanisms. The two distinct autophagic processes were thought to be involved in the breakdown of SG and cell components in the cells of germinated cotyledon.

Show MeSH

Related in: MedlinePlus

A model of autophagic processes in the cotyledon cell of germinated V. mungo seeds. The digestion of storage proteins in germinated cotyledons occurs in cells farthest from the VB. The proform of SH-EP synthesized in the lumen of the ER is packed into the KV at the edge or the middle region of the ER. The KV filled with proSH-EP buds off from the ER, bypasses the Golgi complex, and fuses with the PSV, resulting in the release of proSH-EP into the inside of the PSV. This mass transport of proteinase triggers the breakdown of storage proteins and conversion of the PSV into the LV. α-Amylase is transported to the PSV via the Golgi complex. The acidic vesicle is synthesized de novo (dnV) and fuses with the SG. The transport pathway of dnV to the SG remains open. The possibility that dnVs are derived from the LV by a process of fragmentation of LVs cannot be excluded. An LED area is formed around the SG by fusion of dnVs, and an SG wrapped with LED area is incorporated into the LV by membrane fusion between the LED area around SGs and LVs or engulfment of SGs by LVs. Autophagosome carries mitochondria and cytoplasm to the PSV and/or LV. SGs, mitochondria, and cytoplasm are degraded by hydrolases in LVs. These autophagic processes mediate the change of the cotyledon cells filled with storage materials (shaded cells) into cells in that most cell components are degraded (nonshaded cells). In detached cotyledons, SH-EP is only slightly expressed, and the KV is not formed, resulting in loss of conversion of the PSV to the LV. The acidic vesicle (dnV) is not synthesized in the detached cotyledon cells. AB, autophagic body; AP, autophagosome; dnV, de novo–synthesized acidic vesicle; ER, endoplasmic reticulum; KV, KDEL-tailed cysteine proteinase-accumulating vesicle; LED, low electron density; LV, lytic vacuole; Mt, mitochondrion; PSV, protein storage vacuole; VB, vascular bundle.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2196185&req=5

fig7: A model of autophagic processes in the cotyledon cell of germinated V. mungo seeds. The digestion of storage proteins in germinated cotyledons occurs in cells farthest from the VB. The proform of SH-EP synthesized in the lumen of the ER is packed into the KV at the edge or the middle region of the ER. The KV filled with proSH-EP buds off from the ER, bypasses the Golgi complex, and fuses with the PSV, resulting in the release of proSH-EP into the inside of the PSV. This mass transport of proteinase triggers the breakdown of storage proteins and conversion of the PSV into the LV. α-Amylase is transported to the PSV via the Golgi complex. The acidic vesicle is synthesized de novo (dnV) and fuses with the SG. The transport pathway of dnV to the SG remains open. The possibility that dnVs are derived from the LV by a process of fragmentation of LVs cannot be excluded. An LED area is formed around the SG by fusion of dnVs, and an SG wrapped with LED area is incorporated into the LV by membrane fusion between the LED area around SGs and LVs or engulfment of SGs by LVs. Autophagosome carries mitochondria and cytoplasm to the PSV and/or LV. SGs, mitochondria, and cytoplasm are degraded by hydrolases in LVs. These autophagic processes mediate the change of the cotyledon cells filled with storage materials (shaded cells) into cells in that most cell components are degraded (nonshaded cells). In detached cotyledons, SH-EP is only slightly expressed, and the KV is not formed, resulting in loss of conversion of the PSV to the LV. The acidic vesicle (dnV) is not synthesized in the detached cotyledon cells. AB, autophagic body; AP, autophagosome; dnV, de novo–synthesized acidic vesicle; ER, endoplasmic reticulum; KV, KDEL-tailed cysteine proteinase-accumulating vesicle; LED, low electron density; LV, lytic vacuole; Mt, mitochondrion; PSV, protein storage vacuole; VB, vascular bundle.

Mentions: Autophagic mechanisms have been intensively investigated in yeast, and at least two main autophagic machineries have been identified. First is macroautophagy: cytoplasm and/or mitochondria are sequestered into autophagosomes wrapped with a double membrane, and then the autophagosome fuses with vacuoles (Takeshige et al., 1992; Baba et al., 1994). Second is microautophagy, which was identified in the degradation process of the peroxisome in glucose-treated yeast cells. This autophagic process is proceeded by engulfment of the peroxisome by vacuoles and is called micropexophagy (Tuttle and Dunn, 1995; Sakai et al., 1998). Investigations for autophagy in plants have been mainly conducted with cultured cells, since removal of nutrients in culture medium often induces autophagy in the cultured cells (Chen et al., 1994; Aubert et al., 1996; Moriyasu and Ohsumi, 1996). Therefore, the study of autophagy related to plant development/senescence has been limited. This study indicates that two distinct autophagic processes function in the cotyledon cells of V. mungo seedlings for the degradation of reserves and cellular components. A model of two types of autophagies in a cotyledon cell is presented in Fig. 7 . One autophagic process is the degradation of SGs. In the cotyledon cells, the SG is wrapped with an acidic cell compartment that is possibly built up by the fusion of small acidic vesicle synthesized de novo and subsequently incorporated into the LV that is converted from PSV by the fusion of KV. As for de novo–synthesized vesicles with acidic pH, it is known that small vacuoles are synthesized de novo in the aleurone cells of germinated barley grains (Paris et al., 1996; Swanson et al., 1998; Sansebastiano et al., 2001). The vesicles in the cotyledon cells may correspond to the newly synthesized vacuoles. The degradation of SG in LV was observed by ultrastructural and immunocytochemical analyses of the cotyledon cells. However, we could not clearly identify how wrapped SGs were inserted into the inside of vacuoles. Two possibilities were suggested for the cellular mechanism of the insertion of SGs into LVs. A wrapped SG is further surrounded by several LVs and finally incorporated into the LV through membrane fusion between LVs (Fig. 4 E), or the membrane of the LED region around the SG is fused to that of the LV (Fig. 4 F), resulting in the insertion of SGs into the inside of LVs. It cannot be excluded that both mechanisms function synergistically for effective vacuolar degradation of LED-wrapped SGs. In yeast cells, the peroxisome is wrapped with thin or fragmented vacuoles at the first step of micropexophage, and subsequently the wrapped peroxisome is engulfed into the vacuole (Tuttle and Dunn, 1995; Sakai et al., 1998). Degradation of SGs may be proceeded by a mechanism analogous to micropexophagy, since SG was also wrapped with acidic cell compartments.


Cotyledon cells of Vigna mungo seedlings use at least two distinct autophagic machineries for degradation of starch granules and cellular components.

Toyooka K, Okamoto T, Minamikawa T - J. Cell Biol. (2001)

A model of autophagic processes in the cotyledon cell of germinated V. mungo seeds. The digestion of storage proteins in germinated cotyledons occurs in cells farthest from the VB. The proform of SH-EP synthesized in the lumen of the ER is packed into the KV at the edge or the middle region of the ER. The KV filled with proSH-EP buds off from the ER, bypasses the Golgi complex, and fuses with the PSV, resulting in the release of proSH-EP into the inside of the PSV. This mass transport of proteinase triggers the breakdown of storage proteins and conversion of the PSV into the LV. α-Amylase is transported to the PSV via the Golgi complex. The acidic vesicle is synthesized de novo (dnV) and fuses with the SG. The transport pathway of dnV to the SG remains open. The possibility that dnVs are derived from the LV by a process of fragmentation of LVs cannot be excluded. An LED area is formed around the SG by fusion of dnVs, and an SG wrapped with LED area is incorporated into the LV by membrane fusion between the LED area around SGs and LVs or engulfment of SGs by LVs. Autophagosome carries mitochondria and cytoplasm to the PSV and/or LV. SGs, mitochondria, and cytoplasm are degraded by hydrolases in LVs. These autophagic processes mediate the change of the cotyledon cells filled with storage materials (shaded cells) into cells in that most cell components are degraded (nonshaded cells). In detached cotyledons, SH-EP is only slightly expressed, and the KV is not formed, resulting in loss of conversion of the PSV to the LV. The acidic vesicle (dnV) is not synthesized in the detached cotyledon cells. AB, autophagic body; AP, autophagosome; dnV, de novo–synthesized acidic vesicle; ER, endoplasmic reticulum; KV, KDEL-tailed cysteine proteinase-accumulating vesicle; LED, low electron density; LV, lytic vacuole; Mt, mitochondrion; PSV, protein storage vacuole; VB, vascular bundle.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196185&req=5

fig7: A model of autophagic processes in the cotyledon cell of germinated V. mungo seeds. The digestion of storage proteins in germinated cotyledons occurs in cells farthest from the VB. The proform of SH-EP synthesized in the lumen of the ER is packed into the KV at the edge or the middle region of the ER. The KV filled with proSH-EP buds off from the ER, bypasses the Golgi complex, and fuses with the PSV, resulting in the release of proSH-EP into the inside of the PSV. This mass transport of proteinase triggers the breakdown of storage proteins and conversion of the PSV into the LV. α-Amylase is transported to the PSV via the Golgi complex. The acidic vesicle is synthesized de novo (dnV) and fuses with the SG. The transport pathway of dnV to the SG remains open. The possibility that dnVs are derived from the LV by a process of fragmentation of LVs cannot be excluded. An LED area is formed around the SG by fusion of dnVs, and an SG wrapped with LED area is incorporated into the LV by membrane fusion between the LED area around SGs and LVs or engulfment of SGs by LVs. Autophagosome carries mitochondria and cytoplasm to the PSV and/or LV. SGs, mitochondria, and cytoplasm are degraded by hydrolases in LVs. These autophagic processes mediate the change of the cotyledon cells filled with storage materials (shaded cells) into cells in that most cell components are degraded (nonshaded cells). In detached cotyledons, SH-EP is only slightly expressed, and the KV is not formed, resulting in loss of conversion of the PSV to the LV. The acidic vesicle (dnV) is not synthesized in the detached cotyledon cells. AB, autophagic body; AP, autophagosome; dnV, de novo–synthesized acidic vesicle; ER, endoplasmic reticulum; KV, KDEL-tailed cysteine proteinase-accumulating vesicle; LED, low electron density; LV, lytic vacuole; Mt, mitochondrion; PSV, protein storage vacuole; VB, vascular bundle.
Mentions: Autophagic mechanisms have been intensively investigated in yeast, and at least two main autophagic machineries have been identified. First is macroautophagy: cytoplasm and/or mitochondria are sequestered into autophagosomes wrapped with a double membrane, and then the autophagosome fuses with vacuoles (Takeshige et al., 1992; Baba et al., 1994). Second is microautophagy, which was identified in the degradation process of the peroxisome in glucose-treated yeast cells. This autophagic process is proceeded by engulfment of the peroxisome by vacuoles and is called micropexophagy (Tuttle and Dunn, 1995; Sakai et al., 1998). Investigations for autophagy in plants have been mainly conducted with cultured cells, since removal of nutrients in culture medium often induces autophagy in the cultured cells (Chen et al., 1994; Aubert et al., 1996; Moriyasu and Ohsumi, 1996). Therefore, the study of autophagy related to plant development/senescence has been limited. This study indicates that two distinct autophagic processes function in the cotyledon cells of V. mungo seedlings for the degradation of reserves and cellular components. A model of two types of autophagies in a cotyledon cell is presented in Fig. 7 . One autophagic process is the degradation of SGs. In the cotyledon cells, the SG is wrapped with an acidic cell compartment that is possibly built up by the fusion of small acidic vesicle synthesized de novo and subsequently incorporated into the LV that is converted from PSV by the fusion of KV. As for de novo–synthesized vesicles with acidic pH, it is known that small vacuoles are synthesized de novo in the aleurone cells of germinated barley grains (Paris et al., 1996; Swanson et al., 1998; Sansebastiano et al., 2001). The vesicles in the cotyledon cells may correspond to the newly synthesized vacuoles. The degradation of SG in LV was observed by ultrastructural and immunocytochemical analyses of the cotyledon cells. However, we could not clearly identify how wrapped SGs were inserted into the inside of vacuoles. Two possibilities were suggested for the cellular mechanism of the insertion of SGs into LVs. A wrapped SG is further surrounded by several LVs and finally incorporated into the LV through membrane fusion between LVs (Fig. 4 E), or the membrane of the LED region around the SG is fused to that of the LV (Fig. 4 F), resulting in the insertion of SGs into the inside of LVs. It cannot be excluded that both mechanisms function synergistically for effective vacuolar degradation of LED-wrapped SGs. In yeast cells, the peroxisome is wrapped with thin or fragmented vacuoles at the first step of micropexophage, and subsequently the wrapped peroxisome is engulfed into the vacuole (Tuttle and Dunn, 1995; Sakai et al., 1998). Degradation of SGs may be proceeded by a mechanism analogous to micropexophagy, since SG was also wrapped with acidic cell compartments.

Bottom Line: The results revealed that SG is inserted into LV through autophagic function of LV and subsequently degraded by vacuolar alpha-amylase.When the embryo axes were removed from seeds and the detached cotyledons were incubated, the autophagosome-mediated autophagy was observed, but the autophagic process for the degradation of SG was not detected, suggesting that these two autophagic processes were mediated by different cellular mechanisms.The two distinct autophagic processes were thought to be involved in the breakdown of SG and cell components in the cells of germinated cotyledon.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Tokyo Metropolitan University, Tokyo, 192-0397 Japan.

ABSTRACT
alpha-Amylase is expressed in cotyledons of germinated Vigna mungo seeds and is responsible for the degradation of starch that is stored in the starch granule (SG). Immunocytochemical analysis of the cotyledon cells with anti-alpha-amylase antibody showed that alpha-amylase is transported to protein storage vacuole (PSV) and lytic vacuole (LV), which is converted from PSV by hydrolysis of storage proteins. To observe the insertion/degradation processes of SG into/in the inside of vacuoles, ultrastructural analyses of the cotyledon cells were conducted. The results revealed that SG is inserted into LV through autophagic function of LV and subsequently degraded by vacuolar alpha-amylase. The autophagy for SG was structurally similar to micropexophagy detected in yeast cells. In addition to the autophagic process for SG, autophagosome-mediated autophagy for cytoplasm and mitochondria was detected in the cotyledon cells. When the embryo axes were removed from seeds and the detached cotyledons were incubated, the autophagosome-mediated autophagy was observed, but the autophagic process for the degradation of SG was not detected, suggesting that these two autophagic processes were mediated by different cellular mechanisms. The two distinct autophagic processes were thought to be involved in the breakdown of SG and cell components in the cells of germinated cotyledon.

Show MeSH
Related in: MedlinePlus