Ejercicio 1. Investigar sobre las semejanzas y diferencias entre plantas C3 y C4 e incorporar tres ejemplos de plantas según su metabolismo C3, C4 y CAM. [Photosynthetic metabolism of carbon: type plants C3, C4 and CAM]. [Portuguese ] plantas c3; c4 plants; photosynthese; photosynthesis; carbon; cam pathway; fotosintesis; carbone; voie du metabolisme cam; ciclo cam. From the journal. Las plantas C4 y de metabolismo CAM si tienen ciclo de Calvin, son. reacciones porque no hay fotorrespiración; la fotosíntesis por tanto es más Melvin Calv in y Andrew Benson (): Ciclo C3; marcaje radiactivo en 3-PGA (3C). 2.
Phospho enol pyruvate carboxylase PEPC is distributed in plants and bacteria fotosintesi is not found in fungi and animal cells. Important motifs for enzyme activity and structure are conserved in plant and bacterial PEPCs, with the exception of a phosphorylation domain present at the N terminus of all plant PEPCs reported so far, which is absent in the bacterial enzymes.
Interestingly, this enzyme lacks the phosphorylation domain, hence indicating that it is a bacterial-type PEPC. In contrast, Atppc4 gene has an unusual structure formed by 20 exons. The diversity of plant-type PEPCs in C3, C4, and Crassulacean acid metabolism plants is indicative of the evolutionary success of the fotpsintesis by phosphorylation of this enzyme. In vivo, the active enzyme acts as a homotetramer composed by subunits of to kD O’Leary, This enzyme is widely distributed in plants, algae, and bacteria, but is absent from animals, fotoisntesis, and yeast Lepiniec et al.
All motifs important for enzyme structure and the residues forming the active site are highly conserved fotosintesos PEPCs from plants and bacteria. This is a very important difference because the plant enzyme, but not the bacterial enzyme, is reversibly phosphorylated at this Ser residue by PEPC kinases, which have been recently cloned from Kalanchoe fedtschenkoi and Arabidopsis Hartwell et al.
This regulatory mechanism is also operative in PEPCs from non-photosynthetic tissues as has been shown in developing and germinating cereal grains Osuna et al. In bacteria, PEPC plays an anaplerotic function replenishing intermediates of the citric acid cycle because it is able to recapture CO 2 produced in respiration to generate oxaloacetate, which is readily converted to malate by the action of malate dehydrogenase.
In plants, PEPCs are encoded by small gene families and show differential expression in plant organs that probably reflect specific functions for each gene as has been shown in maize Zea mays ; Kawamura et al. Therefore, it may be distinguished the so-called housekeeping PEPC genes, expressed in tissues with a high metabolic activity, plantaa with an anaplerotic function, and CAM- or C4-specific genes, related to photosynthetic CO 2 fixation, are preferentially expressed in green tissues.
Genes encoding PEPC in plants show a highly conserved structure. This high degree of conservation, and the fact that this enzyme is involved in primary carbon fixation in C4 and CAM photosynthesis, has made PEPC genes very useful molecular markers for the phylogenetic analysis of the different ways of carbon fixation in plants Gehrig et al. These analyses have established that bacterial and plant PEPCs, which have a similar primary structure except for the phosphorylation motif, form two different subgroups Toh et al.
Taking advantage of the sequence of the Arabidopsis genome, we have analyzed the complexity of the PEPC gene family in this plant as a model of dicot, C3 plants. Three of these genes share a high similarity among them and encode typical plant PEPCs. Most notably, it encodes an enzyme without the plxntas motif at the N terminus; that is, a bacterial-type PEPC.
Both PEPC genes, which are not silent, lpantas an unusually high number of introns, suggesting a different evolutionary origin from other known plant PEPC genes.
To obtain the full-length sequence of this gene, a cDNA library was constructed with random primers and mRNA isolated from flowers, the organ showing higher expression of Atppc4 gene see below. AtPPC2 appears annotated with a deletion of 41 residues due to the mis-annotation of sequence that corresponds to exon 9, according to our cDNA data, as intron sequence. Two errors were detected in the annotation of the Atppc4 gene: Exon 3 is actually longer, and the cDNA sequence revealed a new exon exon 10, see belowwhich is annotated as intron sequence.
Atppc1 and Atppc3 code for polypeptides with expected molecular masses of Atppc2 codes for a slightly lower molecular mass polypeptide However, the Atppc4 gene codes fotosintrsis a polypeptide with an expected molecular mass of It may be observed that the insertion responsible of the large size of AtPPC4 residues — do not correspond with the insertions found in the cyanobacterial enzyme Fig.
Sequence alignment of PEPC from different sources. Boxes I to V mark domains important for enzyme catalysis. Surprisingly, no phosphorylation domain was found in the polypeptide deduced from the Atppc4 gene Fig.
This finding, together with the higher level of identity with the E. To our knowledge, this is the first gene encoding a bacterial-type PEPC described in plants; therefore, we tested the presence of this unusual type of PEPC gene in other plants.
The recent release of the rice genome sequence allowed the identification of a similar gene in rice, called Osppc-b here. The polypeptide deduced from Osppc-b included in olantas comparison of Fig.
These are His E. It should be plxntas that the variable residues of this sequence are TV in the polypeptides deduced from Atppc1, Atppc2and Atppc3but SI in the polypeptides deduced from Atppc4 and Osppc-b as in the E.
new concept 4ec7b 7d32f fotosintesis c3 c4 y cam ppt –
Atppc4 is composed of 20 exons, and Osppc-b is composed of 16 exons Fig. Black rectangles, exons; lines, introns. The numbers indicate intron size in nucleotides. Exons and introns in the Arabidopsis genes were deduced by comparing the corresponding cDNA with the gene sequence obtained from the Arabidopsis database http: To establish the phylogenetic relationship of these new plant genes with PEPC genes from different sources, an unrooted phylogenetic tree was constructed using the neighbor-joining method with full-length PEPC amino acid sequences from different bacteria and plants found in accessible databases Fig.
The phylogenetic tree displays two major groups: All these branches were supported by high bootstrap values. The phylogenetic tree shows that both types of plant PEPCs diverged early during the evolution of plants from a common ancestor gene, related to PEPC genes from bacteria such as P. The divergence of Osppc-b and Atppc4 probably reflects the monocot-dicot divergence.
The fact that the bacterial-type PEPC genes found in plants show a different gene structure than the plant-type genes further supports the independent evolution of both types of PEPC genes in plants.
Phylogenetic tree of PEPCs from different sources. The phylogenetic tree was constructed with full-length PEPC amino acid sequences using the neighbor-joining method of the ClustalX version 1. Bootstrap analysis was computed with 1, replicates. Numbers in branches indicate bootstrap values percent.
To distinguish among the different PEPC genes and to detect even low-abundant transcripts, we used the relative quantitative reverse transcriptase RT -PCR technique with gene-specific primers to analyze the expression of this gene family in Arabidopsis. Figure 4A shows a wide distribution of Atppc1 transcripts in all plant organs, though accumulated at higher level in roots.
Atppc2 gene transcripts were also detected in all plant organs but were more abundant in green tissues, such as rosette and stem leaves Fig. In contrast, Atppc3 was almost not expressed in green tissues; these transcripts were more abundant in roots and siliques Fig. An important question addressed in this analysis was to establish whether or not Atppc4the bacterial-type gene, is expressed. The relative quantitative RT-PCR analysis detected Atppc4 transcripts exclusively in siliques, flowers, and, to a lower level, in roots Fig.
It should be mentioned that Atppc4 is expressed at a lower level than the other PEPC genes, as shown by the higher number of cycles needed to obtain the linear range of amplification in the RT-PCR analysis 26 for Atppc1, Atppc2and Atppc3 and 32 for Atppc4. Experiments were repeated at least three times obtaining similar results.
The expected sizes of the bands corresponding to the different transcripts are: Ethidium bromide-stained bands for each gene were quantified with the ScionImage software and compared with the corresponding rRNA band. A search of the databases identified two additional genes in the rice cv japonica group, a root-type PEPC isoform, called Osppc1 here accession no. Both genes are typical plant-type genes. fn
At a lower level, transcripts were also detected in shoots from 5-d-old seedlings and in the scutellum of seeds after 5 d of imbibition and were almost undetectable in the aleurone layer or mature seeds Fig. In contrast, the Osppc2 gene shows a similar level of expression in vegetative tissues, roots, and shoots from 5-d-old seedlings and a lower level of accumulation in seed tissues Fig. This analysis revealed the expression of the bacterial-type Osppc-b gene both in seeds mature seeds and aleurone and scutellum from seeds after 5 d of imbibition and in roots and shoots cm 5-d-old seedlings Fig.
The linear range of amplification for planas Osppc-b gene was of 34 cycles; thus, it also showed a very low level of expression of this gene, as compared with the other Fotosintsis genes from rice, which required a lower number of cycles to obtain the linear range of amplification 26 for Osppc1 and 28 for Osppc2.
Therefore, the RT-PCR analysis shows that the bacterial-type PEPC genes, Atppc4 and Osppc-bare not silent genes, although the level of expression is rather low and, in the case of Arabidopsis, restricted exclusively to some organs.
Differential expression of PEPC genes in rice seeds and seedlings. Rice seeds were allowed to germinate for 5 d and aleurone layer A and scutellum Sc were dissected.
Roots R and shoots Sh were dissected from 5-day-old seedlings. Ethidium bromide-stained bands were quantified with the ScionImage software and compared with the corresponding rRNA band.
Based on the fact that the polypeptides deduced from these genes show slightly more identity with PEPC from Planhas. In addition, these genes have a peculiar gene structure. The average size of introns is larger in Osppc-b than in Atppc4.
This finding is in agreement with the comparison of the intron size in Arabidopsis and rice, based on the sequence and structure of rice chromosome 1, which revealed an average intron size fohosintesis 3. The presence of PEPC in cyanobacteria led to speculate that the plant enzyme might have arisen from an endosymbiotic origin Lepiniec et al. Therefore, based on the phylogenetic analysis and the lower level of similarity with the cyanobacterial PEPCs, a chloroplastidic origin for plant PEPCs is unlikely.
The phylogenetic analysis reported here also suggests that the two types of plant PEPC genes originated very early during the evolution of plants from the fotosintesls ancestor gene. The different number of introns in both types of genes clearly suggests that they have evolved independently since then.
The divergence of Osppc-b and Atppc4 Fig. We have not found PEPC genes after searching the archaeobacteria genomes accessible in public databases.
This finding suggests that the PEPC genes are present in archaeobacteria; however, their sequence must have diverged significantly from PEPC genes characterized thus far. The plant-type PEPC genes, which possess the regulatory mechanism of phosphorylation, form a homogeneous cluster in the phylogenetic tree Fig.
This PEPC is the form that has diverged in different types, allowing the appearance of CAM and C4 photosynthetic metabolisms, which is indicative of the evolutionary success of this mechanism of regulation of PEPC activity. Despite the low level of expression, the bacterial-type genes are not silent and, based on our sequence analysis of Atppc4 cDNA, the introns are correctly processed. These data fotosintesix that the bacterial-type PEPC still plays a role in plant metabolism, but further analysis is required to understand the function of this new type of PEPC in plants.
It is surprising that genes encoding bacterial-type PEPCs in plants have escaped previous analysis of PEPC genes carried out over the years ftosintesis different plants. A possibility, which we think is unlikely, is that these genes are not present in C4 and CAM plants, in which PEPC genes have been more intensively studied. More likely is plantaw, due to the low level of expression of these genes, they are poorly represented in expression libraries and have not been detected.
The fotosintwsis genome sequences of Arabidopsis and rice were ppantas for these unexpected PEPC genes to be identified.
new concept 4ec7b 7d32f fotosintesis c3 c4 y cam ppt
Roots and shoots were dissected from 5-day-old seedlings, and scutellum and aleurone layers were dissected from seeds after 5 d of imbibition.
Five cDNA fragments At to At were obtained ranging in size from to 1, bp with the following primers: For each gene Atppc1Atppc2Atppc3and Atppc4 from Arabidopsis and Osppc1Osppc2and Osppc-b from ricethe linear range of amplification was determined to establish the number of cycles of the PCR reactions.
The pair of primers were: Each pair was used with a mix 1: Control reactions were performed using as template non-reverse transcribed RNA to rule out possible amplification from contaminating genomic DNA.
Ethidium bromide-stained bands were quantified with the ScionImage software, and the amount of mRNA is represented as relative units. Multiple alignment of complete amino acid sequences of PEPC from different plant and bacterial sources accessible from public databases and genome projects was performed with the ClustalX version 1.
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