Thursday, May 21, 2009

Know the Gene Expression

All cells possess the typical number of chromosomes of their species. Therefore, root, epidermis, or pod cells of a soybean plant possess all 40 chromosomes typical of this species. However, not all of the genes are expressed in every cell. For instance, genes that code for chlorophyll production are expressed in the leaves and any other green part of the plant. However they are silenced in the roots, which is the reason these cells do not contain chlorophyll. Gene regulation is a complex process that is affected by a series of factors. A common occurrence in genetic engineering is a lack of expression after a gene has been transformed into an organism. Therefore, an understanding of mechanisms involved with gene expression is critical in genetic transformation.

In bacteria, some genes are activated while others are silenced, depending on the conditions in which these microorganisms are grown. For example, the bacteria Escherichia coli can use two different carbohydrates, lactose and glucose, as energy sources. The bacteria needs to synthesize specific enzymes that catalyze the breakdown of the carbohydrates into energy. The enzymes, like all other proteins, are coded by genes. When E. coli is cultivated in a medium with both glucose and lactose (preferably glucose), it metabolizes. The genes coding for the production of the enzymes that metabolize glucose are thus expressed preferentially. The metabolism of lactose requires an additional enzyme that is only synthesized, or activated, after the medium runs out of glucose and lactose is the only energy source available. This phenomenon is called gene regulation.

Gene expression in more complex organisms is still not completely understood. The complexity of gene regulation is a puzzle in the zygote, a cell formed by the union of sperm and egg cells, in which the genes coding for differing functions have to be activated in a precise and orderly manner. The same genetic information present in the zygote is also present in any other cell in the body, from muscles to skin. Obviously, different genes are activated or expressed in each organ in a different way.

Gene expression is not just a function of where the cell is, but also the result of environmental stimuli. Cells of a floral bud of soybeans differentiate into flowers when the plant is grown during long nights. If the soybean plant is grown during short nights, it continues vegetative growth and does not bloom. Another example of gene regulation occurs with animals, including humans. Testicle and ovary cells do not start the production of sexual hormones until the individual reaches puberty.

Another example of the complexity and importance of gene regulation can be observed in the metamorphosis and development of butterflies and moths. These insects take three forms during their lives: caterpillar, pupa, and adult butterfly or moth. The insect possesses the same genes and DNA during these three different developmental phases. Although the caterpillar, pupa, and adult have the same genes, it is interesting to observe that different genes are expressed in the three developmental phases. In the caterpillar phase, the genes for production of several legs and a stronger mouth capable of chewing leaves are expressed, but not the genes for production of wings. However, the genes for the formation of a delicate mouth apparatus, appropriate for nectar feeding, and genes for the formation of wings are active in the insect's adult phase. The gene expression pattern changes during insect development to allow for the correct progression of its life cycle.

The mechanisms regulating gene expression involve regulatory genes. As opposed to the genes discussed up to this point, these DNA sequences do not code for any protein. Their function is to promote the activation or the silencing of genes.

An important part of gene regulation is the promoters. A promoter is a DNA sequence preceding the gene, which contains regulatory sequences to control the rate of RNA transcription. Promoters control when and in which cells a certain gene is expressed. Through the manipulation of promoters it is possible to induce superexpression, underexpression, or even gene silencing.

Some promoters are constitutive—that is, they induce gene expression continually—whereas others are inducible. Among these, there are some that are chemically inducible, and others are activated by heat, light, or hormones. Some promoters are active in certain tissues and organs, but not in others. In this case, they are considered tissue-specific promoters, as in the case of chlorophyll production. The promoters of the chlorophyll genes are not active in roots, but they are active in the leaves and in all green parts of plants.
Some of the promoters frequently used in genetic engineering of plants include the following:
1) Constitutive
a) UBI from corn
b) 35SCaMV from a cauliflower virus

2) Tissue-specific
a) Phaseolina promoter, a seed-specific promoter from field beans
b) Vicillin promoter, a seed-specific promoter from peas
c) Glutamine promoter, an endosperm-specific promoter from wheat

3) Inducible
a) Rubisco 5S promoter, inducible by light

Aside from promoters, other genetic factors are important in proper gene expression. Although the genetic code is universal, it is also considered degenerate, as more than a single codon codes for a certain amino acid. Different organisms have acquired the preferential use of specific codons for certain amino acids during evolution; this can also have an impact in gene expression. That was the case of the Bt gene from Bacillus thuringiensis introduced in corn. Initially, the expression of that bacterial gene in corn was low; however, when a transgene was reengineered to favor the preferential use of certain codons by corn, gene expression occurred at normal levels.

Several other factors can affect the expression of transgenes, such as the presence of a peptide signal, the site of its integration in the genome, the number of copies integrated, and transgene rearrangements during the integration process. Integration of transgenes in the host genome, in general, happens at random; that is, it can occur in any chromosome of the cell and it can land in any part of the chromosome. However, most of the transgenic varieties have the transgene inserted close to the ends of the chromosome. Multiple copies of the transgene are typically introgressed together.

Tags: Bio Technology, Bio Genetics, Genetic Transformation

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