Dataset Information

Cell type-preferred expression of maize cab-m1: repression in bundle sheath cells and enhancement in mesophyll cells.

ABSTRACT: Different sets of genes for photosynthesis are expressed in mesophyll cells (MCs) and bundle sheath cells (BSCs)--the two adjacent but morphologically and functionally distinct types of photosynthetic cells in leaves of maize and other C4 plants. For example, light-harvesting chlorophyll a/b-binding proteins of photosystem II, which are encoded by a family of cab genes, are 3- to 4-fold more abundant in maize MCs than in BSCs. Each maize cab gene is different from the others in its relative expression in MCs vs. BSCs and in its degree of photoresponsiveness. The gene cab-m1 is positively photoregulated and is highly preferentially expressed in MCs. A 159-bp sequence in the 5' flanking region of this gene (-1026 to -868 relative to the translation start site) is required for MC-preferred expression of a reporter gene in greening maize leaves. Deletion as well as gain-of-function experiments have now shown that all of the sequence information required for MC-preferred expression resides within this mesophyll-specifying region and that cab-m1 is preferentially expressed in MCs because of the presence of two types of sequence elements: one is required for suppressing expression in BSCs and the other for promoting expression in MCs. One of the four cis-acting regions mapped within the mesophyll-specifying region resembles the AT-1 box of some plant gene negative regulatory elements. Various combinations of such MC-specific enhancing and BSC-specific repressing regions could make maize cab gene family members different from one another in their relative expression in MCs vs. BSCs.

SUBMITTER: Bansal KC

PROVIDER: S-EPMC46445 | biostudies-other | 1993 May

REPOSITORIES: biostudies-other

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Cell type-preferred expression of maize cab-m1: repression in bundle sheath cells and enhancement in mesophyll cells.

Bansal K C KC Bogorad L L

Proceedings of the National Academy of Sciences of the United States of America 19930501 9

Different sets of genes for photosynthesis are expressed in mesophyll cells (MCs) and bundle sheath cells (BSCs)--the two adjacent but morphologically and functionally distinct types of photosynthetic cells in leaves of maize and other C4 plants. For example, light-harvesting chlorophyll a/b-binding proteins of photosystem II, which are encoded by a family of cab genes, are 3- to 4-fold more abundant in maize MCs than in BSCs. Each maize cab gene is different from the others in its relative expr ...[more]

PMID: 8483921

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Project description:Chloroplasts of maize leaves differentiate into specific bundle sheath (BS) and mesophyll (M) types to accommodate C(4) photosynthesis. Chloroplasts contain thylakoid and envelope membranes that contain the photosynthetic machineries and transporters but also proteins involved in e.g. protein homeostasis. These chloroplast membranes must be specialized within each cell type to accommodate C(4) photosynthesis and regulate metabolic fluxes and activities. This quantitative study determined the differentiated state of BS and M chloroplast thylakoid and envelope membrane proteomes and their oligomeric states using innovative gel-based and mass spectrometry-based protein quantifications. This included native gels, iTRAQ, and label-free quantification using an LTQ-Orbitrap. Subunits of Photosystems I and II, the cytochrome b(6)f, and ATP synthase complexes showed average BS/M accumulation ratios of 1.6, 0.45, 1.0, and 1.33, respectively, whereas ratios for the light-harvesting complex I and II families were 1.72 and 0.68, respectively. A 1000-kDa BS-specific NAD(P)H dehydrogenase complex with associated proteins of unknown function containing more than 15 proteins was observed; we speculate that this novel complex possibly functions in inorganic carbon concentration when carboxylation rates by ribulose-bisphosphate carboxylase/oxygenase are lower than decarboxylation rates by malic enzyme. Differential accumulation of thylakoid proteases (Egy and DegP), state transition kinases (STN7,8), and Photosystem I and II assembly factors was observed, suggesting that cell-specific photosynthetic electron transport depends on post-translational regulatory mechanisms. BS/M ratios for inner envelope transporters phosphoenolpyruvate/P(i) translocator, Dit1, Dit2, and Mex1 were determined and reflect metabolic fluxes in carbon metabolism. A wide variety of hundreds of other proteins showed differential BS/M accumulation. Mass spectral information and functional annotations are available through the Plant Proteome Database. These data are integrated with previous data, resulting in a model for C(4) photosynthesis, thereby providing new rationales for metabolic engineering of C(4) pathways and targeted analysis of genetic networks that coordinate C(4) differentiation.

Project description:A number of taxa utilize C4 photosynthesis, to limit the impact of photorespiration upon photosynthetic performance. In order to achieve a local elevation of CO2 concentration, maize plants possess two photosynthetic cell types. Rubisco accumulation is restricted to bundle sheath (BS) cells that surround the leaf veins. Carbon fixation occurs initially in adjacent mesophyll (ME) cells. C4 compounds are transported into the BS cells where they are subsequently decarboxylated, releasing CO2. Although the major components of the C4 pathway have been well characterized, less is known about further metabolic partitioning in the maize leaf. Microarray hybridizations have been performed in order to further investigate metabolic differences between BS and ME cell types. BS strands and ME protoplasts were isolated from the leaves of 10 day old maize seedlings by mechanical disruption and enzymatic digestion respectively. To control for differences arising from these different protocols, total leaf (TO) and total leaf stress (ST) samples were also isolated. The ST sample was subjected to the same treatments as the ME sample, with the omission of cell-wall degrading enzymes. Leaves for the TO sample were harvested as for the BS strand sample. An interwoven loop design was used to compare the four treatment groups. A biological group consisted of a growth of plants from which pooled individuals were taken for the four treatments. Six biological replicates (groups) were used. Labeling was performed using the Genisphere Array 900-MPX kit according to the manufacturer's protocol. Post hybridization washes were performed according to the recommendations of the Maize Oligo Array Project. Scan settings were used for detection of moderate to high expression signals (gain ~ 60%. power 90%). Following hybridization with TO cDNA, ~1/3 of features provided signal above twice background and below saturation. A number of taxa utilize C4 photosynthesis, to limit the impact of photorespiration upon photosynthetic performance. In order to achieve a local elevation of CO2 concentration, maize plants possess two photosynthetic cell types. Rubisco accumulation is restricted to bundle sheath (BS) cells that surround the leaf veins. Carbon fixation occurs initially in adjacent mesophyll (ME) cells. C4 compounds are transported into the BS cells where they are subsequently decarboxylated, releasing CO2. Although the major components of the C4 pathway have been well characterized, less is known about further metabolic partitioning in the maize leaf. Microarray hybridizations have been performed in order to further investigate metabolic differences between BS and ME cell types. BS strands and ME protoplasts were isolated from the leaves of 10 day old maize seedlings by mechanical disruption and enzymatic digestion respectively. To control for differences arising from these different protocols, total leaf (TO) and total leaf stress (ST) samples were also isolated. The ST sample was subjected to the same treatments as the ME sample, with the omission of cell-wall degrading enzymes. Leaves for the TO sample were harvested as for the BS strand sample. An interwoven loop design was used to compare the four treatment groups. A biological group consisted of a growth of plants from which pooled individuals were taken for the four treatments. Six biological replicates (groups) were used. Labeling was performed using the Genisphere Array 900-MPX kit according to the manufacturer's protocol. Post hybridization washes were performed according to the recommendations of the Maize Oligo Array Project. Scan settings were used for detection of moderate to high expression signals (gain ~ 60%. power 90%). Following hybridization with TO cDNA, ~1/3 of features provided signal above twice background and below saturation. Keywords: Gene expression profiling of bundle sheath and mesophyll cell types

Project description:During Zea mays (maize) C4 differentiation, mesophyll (M) and bundle sheath (BS) cells accumulate distinct sets of photosynthetic enzymes, with very low photosystem II (PSII) content in BS chloroplasts. Consequently, there is little linear electron transport in the BS and ATP is generated by cyclic electron flow. In contrast, M thylakoids are very similar to those of C3 plants and produce the ATP and NADPH that drive metabolic activities. Regulation of this differentiation process is poorly understood but involves expression and coordination of nuclear and plastid genomes. Here, we identify a recessive allele of the maize Hcf136 homologue that in Arabidopsis thaliana functions as a PSII stability or assembly factor located in the thylakoid lumen. Proteome analysis of the thylakoids and electron microscopy reveal that Zm hcf136 lacks PSII complexes and grana thylakoids in M chloroplasts, consistent with the previously defined Arabidopsis function. Interestingly, hcf136 is also defective in processing the full-length psbB-psbT-psbH-petB-petD polycistron specifically in M chloroplasts. To determine whether the loss of PSII in M cells affects C4 differentiation, we performed cell-type specific transcript analysis of hcf136 and wild-type seedlings. The results indicate that M and BS cells respond uniquely to the loss of PSII, with little overlap in gene expression changes between data sets. These results are discussed in the context of signals that may drive differential gene expression in C4 photosynthesis. To explore the disruption of PSII activity on gene expression, transcript profiles from separated M and BS cells were examined using two-label microarray analysis. Total RNA was isolated from the second leaves of mutant and wild-type silbings. Six biological replicates were used to compare wild-type and mutant transcript profiles in separate M and BS experiments. To maximize biological replication, different seedling pools were used for each of the 12 hybridizations. Microarray experiments and analyses were performed using the Genisphere MPX900 kit and the Maize Array Consortium oligonucleotide platform (GPL5439; GPL5440). Feature intensity values were log-transformed and corrected for local background signal, and a LOWESS procedure (Dudoit et al., 2002) was used to normalize between channels. Features with either low or saturating signal intensity were discarded from further analysis. High expression filtering was less stringent to avoid elimination of previously characterized, high abundance, C4 cell-specific transcripts. After filtering, features that were not assigned an MZ number by the Maize Array Consortium were discarded from further analysis. The moderated t-test (Smyth, 2004) using the R package limma was applied to identify differentially expressed genes. The p-values for each test (gene) were converted to q-values for false discovery rate analysis as described by Storey et al. (2004). To avoid confounding treatment effects associated with direct comparisons of M and BS transcriptomes (Sawers et al., 2007), comparisons were only made using the same cell type across the hcf136 and wild-type sibling genotypes. Bundle sheath (BS) Samples: GSM245063-GSM245164 Mesophyll (M) Samples: GSM245165 - GSM245206

Dataset Information

Cell type-preferred expression of maize cab-m1: repression in bundle sheath cells and enhancement in mesophyll cells.

Publications

Cell type-preferred expression of maize cab-m1: repression in bundle sheath cells and enhancement in mesophyll cells.

Similar Datasets

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