GENE EXPRESSION IN NERVOUS SYSTEM

GENE EXPRESSION IN NERVOUS SYSTEM :

GENE EXPRESSION IN NERVOUS SYSTEM Wajiha Jamal MS Student Biochemistry

GENE EXPRESSION:

GENE EXPRESSION Genes are stretches of DNA that contains the instruction for making or regulating specific proteins. Gene expression is the process by which genes exert their effects on the phenotype of an organism by producing RNA and that synthesized RNA in turn synthesizes protein In order to make a protein, a molecule closely related to DNA called ribonucleic acid (RNA) first copies the code within DNA. Then, protein-manufacturing machinery within the cell scans the RNA, reading the nucleotides in groups of three. These triplets encode 20 distinct amino acids, which are the building blocks for proteins.

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Thus the overall process can be defined as The transcription of a specific DNA base sequence into mRNA and its translation , by a ribosome, into a particular amino acid sequence forming a protein.

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GENE EXPRESSION IN BRAIN

GENE EXPRESSION IN BRAIN :

GENE EXPRESSION IN BRAIN The flow of information from DNA to functional protein is same in the brain as in the other organs Because the cell division has stopped in mature neurons( in majority of regions) the chromosomes no longer duplicate themselves and function only in gene expression

Process of gene expression:

Process of gene expression The whole process of Gene expression occurs in two major stages Transcription Translation In a eukaryotic cell, transcription occurs in the nucleus and translation occurs at ribosomes in the cytoplasm .

TRANSCRIPTION:

TRANSCRIPTION In the process of transcription the synthesis of a single strand RNA complementary to one of the strands of double stranded DNA takes place. For this an enzyme called RNA polymerase II is required which separates the two DNA strands with out making a transcript until it reaches and bonds to a specific sequence called ‘promoter’. Promoters are sequences in DNA that determine the start point and the rate of transcription, they are often composed of smaller sequences called boxes or elements.

Transcription factors:

Transcription factors The RNA polymerase II require the assistance of other protein called transcription factor in order to initiate the synthesis of RNA chain. The transcription factors, rather than the enzyme it selves, are principally responsible for recognizing the promoter and form an appropriate initial complex before the RNA polymerase will bind and initiate transcription. These factors assemble in particular order, beginning with the binding of TFIID to the TATA box, a DNA sequence found just upstream of most eukaryotic RNA polymerase start sites.

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Adenine- and Thymine-rich consensus General Transcription Factor (GTF) binds to TATA box Facilitate the binding of RNA pol II

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All these events lead to the start of transcription in which new nucleotides form a complementary RNA strand using the DNA gene sequence strand as a ‘master’ This process continuous until a specific termination signal on the DNA is reached. In response to this signal RNA polymerase II cease transcription and both RAN polymerase and RNA product in form of primary transcript are released from DNA template.

EUKARYOTIC CELLS MODIFY RNA PRIMARY TRANSCRIPT:

EUKARYOTIC CELLS MODIFY RNA PRIMARY TRANSCRIPT Enzymes in the eukaryotic nucleus modify pre-mRNA before the genetic messages are dispatched to the cytoplasm These modification include Capping Polyadenylation RNA splicing .

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At the 5’ end of the pre-mRNA molecule, a modified form of guanine is added, the 5’ cap . This helps to protect mRNA from hydrolytic enzymes. It also functions as an “attach here” signal for ribosomes. At the 3’ end, an enzyme adds 50 to 250 adenine nucleotides, the poly(A) tail . In addition to inhibiting hydrolysis and facilitating ribosome attachment, the poly(A) tail also seems to facilitate the export of mRNA from the nucleus.

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The most remarkable stage of RNA processing occurs during the removal of a large portion of the RNA molecule during RNA splicing . Most eukaryotic genes and their RNA transcripts have long noncoding stretches of nucleotides. Noncoding segments, introns , lie between coding regions. The final mRNA transcript includes coding regions, exons , that are translated into amino acid sequences, plus the leader and trailer sequences.

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RNA splicing removes introns and joins exons to create an mRNA molecule with a continuous coding sequence .

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RNA splicing appears to have several functions. First, at least some introns contain sequences that control gene activity in some way. Splicing itself may regulate the passage of mRNA from the nucleus to the cytoplasm. One clear benefit of split genes is to enable one gene to encode for more than one polypeptide. Alternative RNA splicing gives rise to two or more different polypeptides, depending on which segments are treated as exons.

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The nervous system provides many examples of the same segment of DNA encoding several polypeptide chains. For example Multiple mRNAs for myelin basic protein Transcription of the gene for myelin basic protein appears to begin and end at single sites to yield a single primary transcript containing seven exons and six introns. Alternative patterns of splicing yield at least five species of mRNA that encodes five polypeptide chains of slightly different sizes. These polypeptides have common sequences at each end and slightly different sequence at the middle.

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No one knows yet whether these five polypeptides have different functions in myelin, but similar alternative splicing of the neural cell adhesion molecule (N-CAM) results in alternative proteins that either contain or do not contain a transmembrane domain, and the two products differ in their manner of association with the cell membrane.

Translation:

Translation The second step of gene expression is the translation, which involves the transfer of the information in mRNA molecules into the sequences of amino acids in polypeptide gene products. That is why translation is also called protein synthesis. The site of protein synthesis are ribosomes which can be found free in the cytoplasm or attached to ER

Requirements for transaltion :

Requirements for transaltion Components Required for Translation Amino acids Transfer RNA (tRNA) Messenger RNA (mRNA), Ribosome Aminoacyl-tRNA synthetases Protein factors Energy source: ATP and GTP

Post translational modification:

Post translational modification Proteins may undergo several modification in order to create their three dimensional structures from linear polypeptide chain. Although the information for a protein’s tertiary structure is encoded in its amino acid sequence, not all proteins fold spontaneously as they are synthesized in the cell. Folding of many proteins is affiliated by the action of specialized proteins that are called “molecular chaperones” that interact with partially folded or improperly folded polypeptides , facilitating correct folding pathways or providing microenvironment in which folding can occur.

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The formation of the tertiary structure also depends on the ionic conditions and pH of the surrounding cytosol, as well as on the concentrations of specific allosteric effectors (small molecules that alter a protein’s conformation and activity).

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Besides the noncovalent modification, the proteins undergo essential covalent modification as well. Covalent modification include Hydroxylation Phosphorylation which may activate or inactivate the protein. This phosphorylation is catalyzed by one of a family of protein kinases and may be reversed by the action of cellular protein phosphatases glycosylation , which targets a protein to become part of a plasma membrane or lysosome or be secreted from the cell. sometimes glycosylation is used to target proteins to specific organelles. and some other covalent modification which may be required for functional activity of a protein like addition of carboxyl group , attachment of lipids besides many proteins are acetylated posttranslationally.

Protein targeting :

Protein targeting Once protein are synthesized in the cytosol and undergone their required modification, the next step is the delivering of protein to its correct location in order for protein to perform its assigned function. The final destination of protein is encoded in its amino acid sequence. A) Major portion of newly synthesized protein remain within the cytosol. Cytosolic proteins include two most abundant groups of protein in a neuron The fibrillar elements that make up the cytoskeleton Nemerous enzymes that catalyze the metabolic reactions of the cell.

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B) Some proteins are synthesized in the cytosol and Actively Imported by the Nucleus, Mitochondria, and Peroxisomes The proteins for these organelles are synthesized on free ribosomes and are imported only after their synthesis is completed. Import into the nucleus takes place through the nuclear pores which require energy of ATP and does not involve the transport through a membrane . The nuclear uptake of these protein depends upon nuclear localization signals in the amino acid sequence of the protein. Proteins that are destined for mitochondria and peroxisomes need to cross the phospholipid bilayer . Thus unlike nuclear proteins, which are imported after they have been folded these polypeptide reach their native conformation only after import into the organelle, which occur soon after the protein is synthesized.

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C. Secretory proteins and Proteins of the Vacuolar Apparatus and Plasmalemma Are Synthesized and Modified in the Endoplasmic Reticulum The translocation of the proteins for these organelles occur during protein synthesis(cotranslational transfer) Besides their mRNA is translated on ribosomes attached to the surface of endoplasmic reticulum. A sequence in the nascent polypeptide induces the attachments of the ribosomes to the rough ER as soon as this portion of the nascent polypeptide chain stars protruding from the ribosomes. This attachment is mediated by a macromolecular complex called the signal recognition particle. Transfer of polypeptide into the ER also requires energy of ATP.

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Some proteins synthesized in the ER remain in the organelle as resident proteins, others are targeted to other compartments of vacuolar apparatus, to the plasmalemma or to extracellular space by secretion. Secretory proteins are processed further in the Golgi Complex and then Exported Proteins exported from the endoplasmic reticulum are carried to the golgi complex in transport vesicles that bud off from the reticulum’s membrane to other intracellular locations or secreted.

Example of a nervous system secretory protein :

Example of a nervous system secretory protein The synthesis of a neuropeptide is very much like the synthesis of any secretory protein made by the cell. First, within the cell nucleus, gene transcription takes place, during which a specific peptide-coding sequence of DNA is used as a template to construct a corresponding strand of messenger RNA. The mRNA then travels to a ribosome, where the process of translation begins. During translation, the sequence of nucleotides that make up the mRNA act as a code to string together a corresponding sequence of amino acids that will eventually become the neuropeptide needed at the terminal. Before this molecule can be transported to the terminal for release into the synaptic cleft, it must be processed in the endoplasmic reticulum (ER), packaged in the golgi apparatus, and transported in storage vesicles down the axon to the terminal.

What makes the difference? :

W hat makes the difference?

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If the process of gene expression is same in the brain as in other organs the question arises that how are the cells of the brain different from cells of the other organs for example the skin cell and the brain cells contain the same set of genetic material but each cell has a unique function within the organism. Both cells for instance carry the genes associated with skin pigmentation, but only the skin cells actually expresses this particular gene and produces the pigment. The answer lies in the fact that only a selected portion of the genetic material or information is transcribed in a given cell to generate mRNAs and eventually proteins. The proteins that are to be transcribed are determined by regulatory DNA –binding proteins (transcription factors)

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These transcription factors also called gene regulatory proteins recognize short stretches of the double –helical DNA of defined sequence and thereby determine which of the thousands of gene in a cell will be transcribed. Hundreds of gene regulatory proteins have been identified each of which has unique features most bind to DNA as homodimers or heterodimers and recognize DNA through one of a small number of structural motifs. Several types of these regulatory proteins identified so far, may have one of three distinctive structural designs helix turn helix (containing three α-helices separated by short turns) zinc finger (which contain zinc atoms bound either to the side chains of four cysteine residues or of two cysteine and two histidine residues) amphipathic helix (which contain α-helices with nonpolar side chains extending from one side, allowing the formation of dimers ). Examples of regulatory proteins that use this structural design are The leucine Zipper motif The helix-Loop-helix motif

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Helix-Turn-Helix Zinc finger Leucine zipper Helix loop helix

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The regulatory proteins are trans acting factors as they are regulatory agents which are not part of the regulated gene(s). These trans-acting factors regulate gene transcription by binding directly or through an intermediate protein to the gene at a particular DNA sequence called a cis -regulatory region. This cis -regulating region is usually located in 5’-flanking promoter region of the gene and is composed of a specific sequences. One class of cis -regulatory elements called enhancers are positioned any where in a gene and consequently are not restricted to the promoter region. These are sequences to which a regulatory protein bind influencing the rate of transcription.

Structural Domains of transcriptional factors :

Structural Domains of transcriptional factors Each transcription factor usually has several structural domains: A DNA-binding domain that recognizes a specific DNA regulatory sequence An effector domain (or transactivating domain) that interacts either with another transcription factor (often forming a homo- or hetro-dimer ) or with a protein of the transcriptional apparatus, thereby increasing or decreasing the rate of transcription; and in some cases, A ligand -binding domain that binds to a small molecule, thereby changing the activity of the factor .

Examples of some transcriptional factors :

Examples of some transcriptional factors A Helix-turn-helix 1. Homeobox (Hox1) 2. POU (Pit-1,Oct-2 ) B Zinc finger 3. Nuclear receptors ( receptors for steroid, thyroid hormones , retinoic acid) C Amphipathic helix 4. Leucine zipper (CREB, fos,jun) 5. Helix-loop-helix (MyoD)

Example CREB:

Example CREB Cyclic AMP, a “second messenger” whose intracellular concentration changes in response to a variety of hormones and transmitters stimulates the phosphorylation (by protein kinase A) of many cellular proteins. One such phosphorylated protein is a ubiquitous transcription factor, with a leucine zipper called CREB (cyclic AMP response element-binding protein). In its phosphorylated form, CREB binds to an 8-base cis -regulatory sequence called CRE (cyclic AMP response element), which is present in several genes whose transcription is increased by cyclic AMP. So increases in cyclic AMP stimulate protein kinase A, which activates CREB, which stimulates transcription.

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Due to regulation of gene expression the cells of nervous system only transcribe those genes whose product (protein) are required for their specialized assigned functions. Thus some of these genes are expressed in all (or the great majority of) neurons and are responsible for those characteristics general to all neuronal cell types (e.g. axon growth, electrical excitability, formation of synaptic junctions). Others are expressed in a subset of neuronal cell types and are responsible for the individual characteristics of particular subpopulations of cells (e.g. neurotransmitter phenotype, enzymes that synthesize neurotransmitter, the pumps that the pumps that exchage ions or recapture neurotranmitter subsatences , and the receptors that transduce physical or biochemical inputs)

Distinctive facts about gene expression in brain :

Distinctive facts about gene expression in brain The gene expression in brain is more complicated but fundamentally the same as in other organs but with some distinctive facts that researchers have found: T he brain expresses more of the total genetic information encoded in DNA than does any other organ in body. About 200,000 distinct mRNA sequence are thought to be expressed 10,20 times more then in the kidney or liver. In part, this diversity results from the greater number and variety of cell types in the brain as compared to cells in the more homogenous body tissues. But many neurobiologist also believe that each of the brain’s 10 11 nerve cells actually expresses a greater amount of its genetic information than does a liver or kidney.

Concluding Remarks:

Concluding Remarks The development of nervous system depends on the expression of particular genes, at particular place and time during development. This spatial and temporal pattern of gene expression is regulated by the hard-wired molecular programs and epigenetic processes

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