2 Gene Expression and Regulation
Walter P. Suza; Faizo Kasule; Philip W. Becraft; and Madan K. Bhattacharyya
Introduction
Every cell in a plant contains the same set of genes and genetic information. Yet different sets of genes are required for the various functions of different cells or tissues, and for plant responses to environmental stimuli or stresses. This is achieved by regulating the activity of genes according to the physiological demands of a particular cell type, developmental stage, or environmental condition. This regulation of gene activity is known as gene expression.
The term expression in genetics can be used in different ways that are sometimes confusing. Typically, the gene is considered “expressed if a gene product is produced.” However, it sometimes occurs that a transcript might be produced but not a protein, or that a protein is produced but in an inactive state. In such cases, although a gene product is made, the biological activity encoded by that gene is absent. For this section, the key point is how the biological activity encoded by a gene is regulated (Figure 1).
The expression of genes in specific plant cells, tissues, and organs, and the timing of this expression, require precise regulation. Expression, or genetic function, can be regulated at any step from transcription, RNA processing, translation, through posttranslational protein modification, as discussed in Chapter 1 under the “Transcription and Gene Structure” section. Regulation can be qualitative, where gene expression is either”on” or “off,” or quantitative, where gene expression levels can be modulated “up” or “down.” Fluctuations in the intensities of external stimuli coupled with changes at the genomic level result in different developmental outcomes or physiological states. Regulation of gene expression at the transcriptional level can occur through chromatin and histone modifications. Also, a gene sequence can be differentially spliced to produce variable-length mRNA products, leading to new protein products with novel functions. Some genes do not encode proteins but short forms of RNAs with regulatory functions, such as the induction of flowering. Finally, protein products can be subjected to modifications such as phosphorylation or dephosphorylation to alter their functions or entirely degraded to turn off a gene.
Transcriptional Gene Regulation
Transcription is the first step in gene expression. Therefore, it makes sense in terms of cellular economy to regulate expression at this level as one of the most critical regulatory points. We already described the involvement of RNA polymerase in the transcription process (see the “Transcription and Gene Structure” section in Chapter 1). However, other protein factors are required. Proteins involved in transcriptional regulation are known as transcription factors. The interaction of transcription factors with specific DNA sequences regulates the process of gene transcription.
The Concept of Differential (Regulated) Gene Expression
As described in the introduction, not every gene is always expressed. When a gene displays different levels of expression in different circumstances, it is known as differential expression. Circumstances that might apply to differential expression include, but are not limited to, different plant tissues (root vs. leaf), different developmental stages (germination vs. reproductive development), or in response to different environmental stimuli (cold stress or pathogen attack).
The term differential expression can also be used to compare the expression of different genes. If two genes show different expression patterns (among plant tissues or in response to environmental stimuli), they are considered differentially expressed, whereas genes with similar expression patterns would be regarded as co-expressed.
Promoters
As mentioned, and discussed in Chapter 1, transcription is regulated through the interactions of proteins, transcription factors, with specific DNA sequences. Most regulatory DNA sequences governing gene transcription are located on the 5′ border of the transcribed region. This region is called the gene promoter. Promoters contain a core required to bind the “basal transcriptional machinery”, including RNA polymerase. The “TATA” box, with a consensus sequence TATAA, is located within the core promoter, usually 25-30 nucleotides upstream of the transcription initiation site. Promoters also contain regulatory sequences that determine when, where, and to what level genes are transcribed. Promoters can vary in length from one hundred to 1000 nucleotides (100–1,000 base pairs).
The complete promoter sequences of different genes expressed similarly may differ. However, such promoters often contain short sequence “motifs” that are similar, referred to as cis elements. Early work (Benfy and Chua, 1990) to understand the function of different promoter elements in regulating gene expression in plant cells and tissues revealed that various combinations of cis elements can be interpreted by the cell and control gene expression. Sometimes, cis elements promote gene transcription, and other times restrict gene transcription in particular cells and tissues.
Promoter analysis is facilitated by using reporter genes. The reporter gene produces easily identifiable and quantifiable effects, which can be used to determine the function of a regulatory region of another gene (promoter, promoter elements, enhancers) in cells, tissues, or organs. Such analyses are critical to crop biotechnology, where targeted gene expression in particular tissues is often desirable. To test whether a promoter is effective in conferring the expression of a gene to a particular tissue, scientists fuse the putative promoter to a reporter gene and introduce the promoter-gene fusion into plants (He et al., 2020). An example of a reporter gene is RUBY (Figure 2), which is a fusion of three genes: (1) CYP76AD1 encodes a cytochrome P450 enzyme that catalyzes the conversion of tyrosine to L-3,4-dihydroxyphenylalanine (L-DOPA) and oxygenation of L-DOPA to cyclo-DOPA; (2) the next enzyme in the reporter gene fusion, DODA ( L-DOPA 4,5-dioxygenase), makes an enzyme that cleaves L-DOPA to form betalamic acid. The enzyme converts betalamic acid and cyclo-DOPA spontaneously to form betanidin, (3) glucosyl transferase (GT), which glycosylates betanidin to produce betalain, a natural red pigment. Because betalain has a very bright red color, which is vividly visible to the naked eye, the RUBY system allows rapid and non-destructive monitoring of plant transformation and gene expression without the need for specialized equipment, expensive chemicals, or invasive treatments (He et al., 2020). The three gene fusions are separated by P2A peptides (Figure 2).
The ectopic expression of RUBY leads to visible pigment accumulation in plant tissues in many plants, like Arabidopsis, tomato, eggplant, carrot, and maize, among others. Strong promoters like Cauliflower Mosaic Virus 35S (CaMV35S) and Ubiquitin have been used to drive the constitutive RUBY expression throughout Arabidopsis and maize. In contrast, seed-specific promoters such as the At2S3 from Arabidopsis restrict RUBY expression to seeds .
Figure 3 shows an example of RUBY expression in various maize tissues using the P35S promoter and Arabidopsis HSP18.2 terminator. Using this constitutive 35S promoter to drive RUBY expression, betalain accumulation was observed from early stages and across the different tissues and developmental stages in maize (Figure 3). This indicates that RUBY serves as an excellent marker for maize transformation (Lee et al., 2023).
Enhancers
Enhancers are DNA sequences that increase the rate of transcription of a gene when they are present, although they cannot cause transcription to occur when alone. Enhancers are usually position and orientation-independent. Although normally located upstream of the promoter, enhancers can also be located on the 3′ region of the gene or even within the coding region. They can increase the transcription when added to genes they are usually not associated with. This is a useful property for biotechnology, allowing promoters to be manipulated for increased levels of transcriptional regulation. Some enhancers, referred to as constitutive, always function in all cells and tissues. Other enhancers function in specific tissues at specific developmental stages. Some enhancers are active only in response to environmental signals. The AACCA enhancer on the promoter of the soybean β-conglycinin gene encoding a seed storage protein functions specifically in seeds. Enhancers interact with specific nuclear proteins involved in transcription. For example, enhancers might facilitate the binding of transcription factors and direct those factors along the DNA strand in the direction of the promoter. Alternatively, enhancers may facilitate changes in DNA structure, such as modification of the chromatin structure.
The counterpart of an enhancer is a silencer. Silencers have all the properties described for enhancers, except they dampen, or decrease, the transcription levels controlled by a promoter.
Transcription Factors
RNA polymerase binds the promoter at the TATA box and drives gene transcription in cooperation with other proteins called transcription factors. Transcription factors interact with RNA polymerase to facilitate its binding to the promoter and regulate its activity. Some transcription factors, known as “basal transcription factors,” are fundamental to the RNA polymerase binding and function, and are expressed in all living cells.
Other transcription factors bind to cis-regulatory DNA sequences of promoters, enhancers, or silencers. These proteins interact in complex ways with the basal transcription machinery to regulate the activity of RNA polymerase, and therefore gene transcription. Thus, transcription factors regulate when or where individual genes are expressed, and to what level.
Transcription factors can function as either positive or negative regulators. That is, they can either function to induce (increase) gene transcription or to repress it. The consequence of many transcription factors depends on their interactions with other proteins. Two factors together may be required for gene activity, and the exclusion of one of the factors in space and time offers a mechanism for differential gene expression. Some transcription factors might function as a positive regulator in one context but as a negative regulator in another, depending on what other cis elements and/or transcription factors might be present.
- About 6% of the 30,000 genes in the Arabidopsis genome encode transcription factors. Most of these genes occur in families. For more information read: Arabidopsis Transcription Factors: Genome-Wide Comparative Analysis Among Eukaryotes
- Transcription factor biotechnology is gaining interest in efforts to solve crop production challenges in unfavorable environments. For more information read: Plant nuclear factor Y (NF-Y) B subunits confer drought tolerance and lead to improved corn yields on water-limited acres
- Read: Regulating the Regulators: The Future Prospects for Transcription-Factor-Based Agricultural Biotechnology Products
Epigenetic Regulation
Chromatin Structure and Histone Modification
At the molecular level, chromatin is a product of ordered and tight packaging of the double-stranded DNA around nucleosomes (a core of histone proteins) and an association with additional proteins.
Transcriptional regulation often involves modification of chromatin structure mediated by posttranslational regulation (changes in acetylation and methylation) of histones. Histone acetylation involves the addition of acetyl groups, and when histones are heavily acetylated, the DNA is less tightly associated with them. This often correlates with increased transcriptional activity of specific genes. The idea is that when DNA is loosely associated with histones, it is more accessible to transcription factors that require interaction with the DNA to initiate transcription. Consequently, histone deacetylation (removal of acetyl groups) by histone deacetylase enzymes (FLD, p462) stabilizes nucleosomes and represses transcription. On the other hand, histone acetylation by histone acetyltransferase destabilizes nucleosomes and promotes transcription.
As described in the text, another form of histone modification is the addition of methyl groups to histone proteins, which similarly regulate chromatin condensation or decondensation.
DNA Methylation
The nucleotide bases of DNA can be modified by the attachment of methyl groups at various locations. The addition of these methyl groups happens after the DNA has been synthesized and is controlled by enzymes that add methyl moieties to specific regions of DNA. The most common modified nucleotide base is C5 methylcytosine (m5C) due to the activity of the enzyme DNA (cytosine-5) methyltransferase (MET). MET recognizes the unmethylated newly replicated DNA strand and incorporates a methyl group if the template strand was methylated (hemimethylated).
Methylation status is correlated with gene expression (Figure 4) as demonstrated by the low level of methylation in regions of the genome undergoing active transcription.
Researchers use the DNase sensitivity assay to determine the genomic regions undergoing active transcription (Chapter 1). DNAse-sensitive regions contain lower methylation than regions that are resistant to the enzyme. The specific methylated DNA sites can be determined using restriction enzymes that recognize the same sequence but differ in their ability to digest DNA if their target site is methylated. For example, the enzyme HpaII cleaves CCGG only if not methylated. However, MspI cleaves CCGG if it is unmethylated or methylated. Figure 5 shows an example of a DNA sequence containing stretches of CCGG that are variably methylated. Using the example of SNP analysis in Plant Breeding Methods, an experiment can be designed where DNA is isolated and subjected to the two restriction enzymes. The products of the two enzymes can be visualized on a gel as shown in Figure 5.
Studies have shown that altering a plant’s overall DNA methylation status can affect growth and development. For example, cold treatment (vernalization) induces flowering in biannuals such as Arabidopsis and winter wheat and lowers the level of methylation of specific genes. Also, treating plants with the drug 5-azacytidine prevents methylation at the 5’ position of cytosine and stimulates flowering. Recent studies also suggest a relationship between epigenetic gene regulation and heterosis, with hybrids showing higher global transcription levels, higher histone acetylation levels, and lower DNA methylation levels.
RNA-Level Regulation
Regulation of gene expression occurs at many levels, including post-transcriptionally (Figure 6). From the expression level for a given gene, a critical factor is the level of fully processed, mature mRNA. When we consider the level of the mature form, a particular transcript at any given time reflects the steady state balance between synthesis, processing, and degradation. The post-transcriptional regulation of RNA occurs through several mechanisms. Figure 6 summarizes how post-transcriptional regulation is essential for plant cells to respond to stress.
Alternative Splicing
Alternative splicing is the mechanism of alternatively processing pre-mRNAs to generate mRNAs with different exon combinations. In animals, many genes are spliced in alternative ways, leading to multiple proteins from a single gene. According to some estimates, over half of all human genes produce alternately spliced transcripts. Interestingly, as described in the textbook, the sexual differences between male and female flies are determined by alternate RNA splicing and the different resulting proteins. One spliced form of the Tra transcript produces male flies; an alternatively spliced Tra transcript results in female flies.
Over 20% of multi-exon gene transcripts undergo alternative splicing in rice and Arabidopsis.
Examples of alternative splicing in plants include the maize pyruvate orthophosphate dikinase (PPDK) gene involved in C4 photosynthesis (Sheen, 1991). Alternative splicing of the PPDK gene results in a protein with either a short stretch of amino acids that keeps it in the cytoplasm or another that targets it to the chloroplast.
Another example is the Jasmonate Zim-Domain (JAZ) genes involved in the signaling pathway of jasmonic acid. Jasmonic acid is required for insect and pathogen defense and reproductive development. In Arabidopsis, the JAZ10 gene produces alternately spliced transcripts with three splice variants (Chung and Howe, 2009). When one of the splice-variants, JAZ10.4, is overexpressed, it causes male sterility by producing a protein that interferes with JA signaling in the flower, preventing the development of stamens.
RNA Stability and Degradation
The control of mRNA degradation rate, or turnover, is an important regulatory mechanism in gene expression. As mentioned, the level of a particular transcript at any given time reflects the steady-state balance between synthesis and degradation. Thus, even though a gene may be transcribed at a high rate, a high rate of RNA turnover could effectively shut off the gene. The stability of mRNA can be regulated globally to affect all or most transcripts, or it can be very specific to a particular mRNA. Regulated mRNA turnover may be significant in plants since plants cannot move to avoid harsh environmental conditions, and stresses are known to induce specific changes in RNA stability (Nakaminami et al., 2011). Other factors, such as light and plant hormones, are also known to regulate mRNA stability.
The degradation of mRNA is catalyzed by enzymes called ribonucleases. Ribonucleases include exonucleases, which degrade only from one end of the transcript, and endonucleases, which can attack the mRNA molecule internally (Figure 7).
Therefore, controlling ribonuclease access to the mRNA substrate is an essential regulatory mechanism in gene expression. This control is achieved through various mechanisms, including controlling the amount of nucleases present, regulating the activity of the nuclease, sequestering the nuclease to specific cellular locations to restrict its access to RNA, and controlling the stability of the mRNA by decreasing accessibility to nucleases.
Small RNAs and RNA Interference
RNA interference (RNAi) refers to RNA-mediated regulation of gene expression. It is a natural mechanism for moderating gene expression in the cells of all eukaryotic organisms, from plants to humans. RNAi usually results in decreased expression or silencing of a target gene through several different mechanisms, including transcriptional silencing, translational silencing, or mRNA degradation. In plants, mRNA degradation is perhaps the most common mechanism of RNAi. These processes are mediated by small RNAs that target mRNA or chromosomal genes through sequence homology. Several small regulatory RNAs exist, including microRNAs (miRNAs) and small interfering RNAs (siRNAs). The formation of siRNAs or miRNAs is initiated by the formation of double-stranded RNA (dsRNA), which is then processed by Dicer endonucleases to form the small RNAs of 21-24 nucleotides. This RNA then associates with a protein complex called RISC and targets cellular mRNAs with homologous sequences.
RNAi is an important mechanism by which endogenous genes are regulated. It commonly functions to refine the pattern of gene expression. For example, a maize gene called tasselseed4 (ts4) codes for a microRNA called miR172. This microRNA is required to target the degradation of mRNAs for several related genes. Those target mRNAs are transcribed in male flowers but usually do not accumulate there. However, when the ts4 gene is mutant, miR172 is not produced. The target mRNAs and their encoded proteins accumulate in male flowers, causing them to develop female characteristics, and hence, seeds form on tassels (Chuck et al., 2007).
RNAi can be accomplished more efficiently by expressing a portion of the target gene that has been engineered as an inverted repeat in transgenic crop plants. Following transcription of this engineered gene, the RNA molecules form a hairpin structure that is then cleaved into small fragments of double-stranded RNA, which interfere with the accumulation and function of the endogenous mRNA molecules of the target gene.
Discovery of RNAi
Although researchers working with worms received full credit for the discovery of RNAi (Fire et al., 1998), related phenomena had been previously observed in plants (Napoli et al., 1990). In the early 1990s, Richard Jorgensen and co-workers, working with petunia, explored ways to increase flower pigmentation through transgenic approaches. Surprisingly, Jorgensen and his team observed that overexpression of transgenes that were intended to increase pigmentation in petunia did not deepen pigmentation. Instead, they observed flowers that showed variegated pigmentation, with some lacking pigments altogether (Figure 8). The researchers concluded that the transgenes and endogenous sequences homologous to the transgenes must have been inactivated, a process they coined co-suppression (Napoli et al., 1998). Other laboratories later found that plants respond to RNA viruses by destroying viral RNAs to prevent viral genes from being expressed inside plant cells, and cause disease. Co-suppression and viral RNA targeting processes in plant cells are referred to as post-transcriptional gene silencing (PTGS).
Post-transcriptional Gene Silencing
Many RNA and DNA viruses stimulate a process known as posttranscriptional gene silencing (PTGS) immediately after infecting a plant. PTGS is proposed as an anti-viral defense mechanism in plants. The most crucial trigger in PTGS is the production of double-stranded RNA by the virus during viral multiplication. Viral RNAs are recognized as foreign and are targeted for degradation in infected cells through RNAi.
It is now well established that PTGS in plants is an RNA-degradation mechanism that is like RNA interference (RNAi) in animals (Figure 9). In plants, mRNA degradation is perhaps the most common mechanism of RNAi. As mentioned above, RNAi usually results in decreased expression or silencing of a target gene The formation of siRNAs) or miRNAs) is initiated by the formation of dsRNA molecules which are processed by dicer endonucleases to form the small RNAs of 21-24 nucleotides. Small RNAs then associate with a protein complex called RISC (RNAi silencing complex) and target cellular mRNAs with homologous sequences to result in reduced protein expression (Figure 8). miRNAs act at the post-transcriptional level leading to translational repression, degradation of the endogenous transcripts and silencing of genes through epigenetic means.
Translational Regulation
As described earlier, for most genes, the protein product of the gene performs the biological function. Therefore, regulating the amount of protein production effectively governs gene expression. There are several known mechanisms by which the process of translation is regulated. A detailed consideration of these mechanisms is beyond the scope of this course, but generally, it is important to bear in mind their importance. For example, under certain stress conditions, “normal” translation is halted and only mRNAs related to stress tolerance are selectively permitted to be translated into proteins. This selective translation is important to allow plants to respond to stress quickly and to conserve energy under stress conditions.
Translation of mRNAs can play an important role in determining their overall stability. Mutations that add premature stop codons often lead to rapid mRNA degradation. Ribonucleases (see RNA Stability and Degradation section in Chapter 2) or other factors involved in degradation may recognize the number or spacing of ribosomes on an mRNA and degrade those that are not produced properly. This may help prevent the synthesis of proteins with incorrect functions, which will negatively impact cellular processes. During transcription, errors could add or omit nucleotides, altering the proper codon sequence and creating mutant proteins (Chapter 3).
Protein-Level Regulation
After translation, proteins are subject to various modifications that can regulate their activity. There are many ways by which proteins can potentially be modified, and thereby regulated, and only a few of the basic mechanisms will be considered here.
Common Types of Post-Translational Modifications
The first mechanism is covalent modification by the addition of various chemical groups. A wide variety of groups can be involved, including small organic groups (methylation, acetylation), lipids (myristoylation, farnesylation, palmitylation), carbohydrates (glycosylation, glucosylation), small proteins (ubiquitination, sumoylation), and inorganic molecules (phosphorylation, sulfation). Such covalent modifications are generally accomplished by the activity of enzymes specialized to perform these modifications. Many of these modifications are also reversible; these groups can be added to a protein and subsequently removed. Phosphorylation is a particularly noteworthy reversible modification that is common in the regulation of many proteins. Enzymes that add phosphate groups to other proteins are called protein kinases, and those that remove phosphates are called phosphatases. As such, protein kinases and phosphatases are central to many cellular regulatory systems.
A second common mechanism of protein modification is through proteolytic cleavage. Proteolysis occurs as part of the general turnover of cellular proteins, which is required to eliminate damaged proteins and recycle amino acids. Proteolysis is important for processing certain proteins, for example, in removing the signal peptide of proteins targeted to specific cellular compartments. It can also occur in a highly specific manner, whereby particular proteins are targeted for degradation or cleavage at a specific site within the protein. Proteolytic modifications are non-reversible.
Proteins can undergo modification through complex formation. Such complexes can occur among proteins or between a protein and a cofactor.
Proteins can also be modified according to the conditions of the cellular environment. The redox state can result in oxidation or reduction of proteins, particularly sulfhydryl side groups. Cellular pH can affect the charge of ionizable side groups.
Common Types of Protein Regulation
All the above-mentioned types of protein modification can alter protein conformations and thus have regulatory consequences on protein function or activity. Protein regulation is highly complex, and there are a myriad of different ways in which this occurs. Again, we will briefly consider a few of the more common mechanisms.
One major way protein modification can regulate protein function is by altering its activity. This can be true for many types of proteins, including enzymes, transcription factors, signaling proteins, and structural proteins—for example, a group of transcriptional regulators known as WRKY transcription factors function in several stress responses. WRKY8 is important for pathogen defense. Upon infection, WRKY8 becomes phosphorylated, which activates its DNA-binding activity and results in the activation of genes involved in pathogen defense (Ishihama et al., 2010).
Cells are compartmentalized (Figure 10) into several membrane-bound organelles, including the nucleus, chloroplasts, mitochondria, peroxisomes, endoplasmic reticulum (ER), golgi, and vacuoles. Each of these compartments performs unique metabolic functions that require a set of proteins.
Compartmentalization is regulated for some proteins. For example, upon exposure to light, the phytochrome protein moves into the nucleus, affecting light-regulated gene expression.
Proteolytic processing is involved in several important regulatory processes. Many proteins are synthesized in an inactive form that requires proteolytic cleavage for activation. The full-length translation product before processing is often called the precursor, or a preprotein. As mentioned, cells contain membrane-bound compartments. Since membranes are impermeable to most proteins, an active mechanism is necessary to move a protein across a membrane. For proper delivery to their organellar destinations, specific amino acid sequences, called target signals, must be present in a protein (to serve as an “address”). For example, entry into the chloroplast is achieved by the presence of a target signal called the transit peptide. This is at the amino-terminal end of the protein and is proteolytically cleaved during import.
Another example of proteolytic processing is seen in a plant defense response in members of the Solanaceae (for example, tomato and potato). An 18-amino acid peptide hormone called systemin is secreted by plant cells damaged by insects or mechanical wounding. Systemin production by wounded cells is required to induce the synthesis of proteins involved in defense. Systemin induces defense responses in wounded cells and throughout the plant. Analogous to animal peptide hormones (e.g., insulin), systemin is initially synthesized as a much larger (200 amino acids) precursor called pro-systemin. Pro-systemin is inactive; however, upon wounding, it undergoes proteolytic cleavage to produce activated systemin.
Targeted Protein Degradation
The amount of protein present in a cell or tissue is determined by both its rate of synthesis and its rate of degradation. Therefore, protein degradation is an important mechanism by which the plant can regulate biological activity (i.e., a genetic function). For example, one way to shut down a metabolic pathway is by degrading one of the key enzymes controlling the rate of the entire pathway. Therefore, protein degradation is an essential component of gene regulation to meet cellular demands for growth, development, and defense.
Protein degradation must be carefully controlled to fine-tune gene expression to allow plants to adapt to new environmental conditions. Often, cells will adopt several complex mechanisms for proteolytic degradation of proteins. Enzymes that cleave or degrade proteins are referred to as proteases. For example, plant vacuoles (Figure 8) are rich in proteases that function similarly in protein degradation as lysosomes in animal cells. Protease activity must be tightly regulated to prevent accidental degradation of essential proteins. Sequestering proteases in particular organelles, like the vacuole, separates them from other organelles and is one means to control their activity.
An important mechanism by which specific proteins are targeted for degradation is through ubiquitin-mediated proteasomal degradation. The proteasome is a large complex of multiple protein subunits with protease activity to degrade proteins. Proteins get marked for proteasomal degradation with a small protein called ubiquitin. Ubiquitin is covalently attached to specific proteins in response to environmental or developmental signals. This allows plants to quickly adapt to changing conditions by eliminating proteins whose functions are not advantageous under the new conditions. For example, the photoreceptor phytochrome mentioned above becomes targeted for degradation when light is no longer available. This allows plants to change their physiological functions, going from daylight to night conditions. Protein degradation by the proteasome system is also an important regulatory mechanism for plant hormone signaling, for example, the signaling of the defense hormone jasmonic acid and the growth hormone gibberellic acid.
Proteins have lifetimes that range from a few minutes to weeks or more. Cells continuously make proteins from mRNA molecules and break them down into amino acids. One of the functions of protein degradation is to eliminate aberrant or damaged proteins, which could harm the cell. The second function is to facilitate the recycling of amino acids. For example, most of the amino acids required for seedling growth are derived from the degradation of seed storage proteins. Conversely, in annual crop plants, many amino acids in seed storage proteins are derived from proteins degraded in leaves and other plant parts during senescence.
Chapter Summary
The expression of genes in specific plant cells, tissues, and organs, and the timing of this expression, require a precise level of regulation. A single promoter may not be sufficient to regulate the expression of such gene(s) in space and time. Therefore, coding regions with the same function may have different promoters, and such genes are referred to as differentially regulated. Most regulatory sequences governing gene expression are located on the 5′ border of the coding region. Transcription is often initiated between 20 and 60 nucleotides upstream of the ATG start site. Enhancers are usually position- and orientation-independent. Although they are normally located upstream of the promoter, they can also be located on the 3′ region of the gene or even within the coding region. An enhancer can increase transcription levels when added to genes they are not normally associated with. RNA polymerase binds the promoter at the TATA box and drives gene transcription in cooperation with other proteins. The proteins that interact with RNA polymerase bind to regulatory sequences upstream and downstream of the transcription site. Transcription factors can play a regulatory role by determining where individual genes are expressed. Transcriptional regulation often involves chromatin modification by changes in acetylation and methylation of histone. The nucleotide bases of DNA can be modified by the attachment of methyl groups at various locations. Methylation status is correlated with gene expression. Alternative splicing describes an alternative mechanism of pre-mRNA processing to generate mRNAs with different exon combinations. The control of mRNA degradation rate, or turnover, is an important regulatory mechanism in gene expression. RNA interference (RNAi) is a post-transcriptional process involving the degradation of mRNA initiated by the formation of double-stranded RNA (dsRNA) of the target mRNA. Translation of mRNAs can play an important role in determining their overall stability. Mutations that add premature stop codons often lead to rapid mRNA degradation. Protein degradation is an essential component of gene regulation to meet cellular demands for growth, development, and defense. The plant can alter the activity of a metabolic pathway by degrading one of the key enzymes controlling the rate of the entire pathway.
References
Ali N, Datta SK, Datta K. RNA interference in designing transgenic crops. GM Crops. 2010 Jul-Sep;1(4):207-13. doi: 10.4161/gmcr.1.4.13344. PMID: 21844675.
Chuck, G., Cigan, A.M., Saeteurn, K., and Hake, S. 2007. The heterochronic maize mutant Corngrass 1 results from overexpression of a tandem microRNA. Nat. Genet. 39:544-549. Available at http://www.nature.com/ng/journal/v39/n4/pdf/ng2001.pdf
Chung HS, Howe GA. A critical role for the TIFY motif in repression of jasmonate signaling by a stabilized splice variant of the JASMONATE ZIM-domain protein JAZ10 in Arabidopsis. Plant Cell. 2009 Jan;21(1):131-45. doi: 10.1105/tpc. 108.064097. Epub 2009 Jan 16. PMID: 19151223; PMCID: PMC2648087.
He, Y., Zhang, T., Sun, H., Zhan, H., & Zhao, Y. (2020). A reporter for noninvasively monitoring gene expression and plant transformation. Horticulture Research, 7 (1), 1-6. https://doi.org/10.1038/s41438-020-00390-1
Ishihama, N., Yamada, R., Yoshioka, M., Katou, S., and Yoshioka, H. 2011. Phosphorylation of the Nicotiana benthamiana WRKY8 Transcription Factor by MAPK Functions in the Defense Response. The Plant Cell. 23: 1153-1170. Available at http://www.plantcell.org/content/23/3/1153.full.pdf
Lee, K., Kang, M., Ji, Q., Grosic, S., & Wang, K. (2023). New T-DNA binary vectors with NptII selection and RUBY reporter for efficient maize transformation and targeted mutagenesis. Plant Physiology, 192 (4), 2598-2603. https://doi.org/10.1093/plphys/kiad231
Nakaminami, K., Matsui, A., Shinozaki, K., and Seki, M. 2012. RNA regulation in plant abiotic stress responses. Biochimica et Biophysica Acta (BBA). Gene Regulatory Mechanisms. Available at http://www.sciencedirect.com/science/article/pii/S1874939911001386
Nelson, D.E., Repetti, P.P., Adams, T.R., Creelman, R.A., Wu, J., Warner, D.C., Anstrom, D.C., Bensen, R.J., Castiglioni, P.P., Donnarummo, M.G., Hinchey, B.S., Kumimoto, R.W., Maszle, D.R., Canales, R.D., Krolikowski, K.A., Dotson, S.B., Gutterson, N., Ratcliffe, O.J. and Heard, J.E. 2007. Plant nuclear factor Y (NF-Y) B subunits confer drought tolerance and lead to improved corn yields on water-limited acres. Proc. Natl. Acad. Sci. USA: 16450-16455. Available at http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2034233/pdf/zpq16450.pdf
Sheen, J., 1991. Molecular mechanisms underlying the differential expression of maize pyruvate orthophosphate dikinase genes. The Plant Cell 3:225-245. Available at http://www.plantcell.org/content/3/3/225.full.pdf
