NE 102 - Second Midterm Block Flashcards

Transcription

A complex process that produces RNA strands complementary to parts of a DNA strand

RNA

A “decoder” of genetic information that can be used in the cytosol; Includes mRNA, tRNA, rRNA, miRNA, all with a purpose to regulate expression of proteins in a cell

mRNA

codes for proteins

ribosomal RNAs

form the core of the ribosome’s structure and catalyze protein synthesis

tRNAs

serve as adaptors between mRNA and amino acids during protein synthesis

RNA orientation

RNA molecules are oriented 5’-3’ and includes phosphodiester bonds; RNA is single stranded, allowing for flexible folding, multiple functions, and can make interactions via H bonds

RNA Folding

Bases find complementary sequences within the same molecule of RNA

RNA Polymerase

Transcribes genetic information into RNA; A protein complex responsible for synthesis of RNA from a DNA template during transcription. Amount of RNA pol in a cell’s nucleus is very high.

RNA Polymerase I and III

Most rRNA genes, tRNA genes, small RNAs

RNA Polymerase II

All protein-coding genes, miRNA genes, and genes for noncoding RNAs (Ex. those in spliceosomes)

RNA Synthesis

No primer or helicase needed, only copies selected regions called genes

Effective Transcription

RNA must recognize gene, the strand it is on, the start point, and needs to be tightly bound to DNA

Promoter

A DNA sequence needed for RNA pol to recognize the gene that needs to be transcribed. Found on the strand that will NOT be copied, located near 5’ end. RNA pol will attach to DNA in a region close to the promoter. Acts as the “TATA box,” easier to unwind.

TATA Box

A conserved DNA sequence in the promoter region that facilitates the binding of RNA polymerase and is essential for the initiation of transcription.

Transcription in Eukaryotic Cells

Transcription begins with “bending” DNA structures through TFs. Specific DNA sequences allow the beginning and the end of transcription. Transcription occurs in the nucleus and requires several processing steps, including the addition of a 5' cap and poly-A tail.

Transcription Factors

TFs bind to promoter and helps RNA pol bind to DNA. Helps prepare DNA and RNA pol. Optimizes RNA pol attachment and activity.

TBP - TATA Binding Protein

Changes conformation of the double helix; TBP is a key subunit of the general transcription factor TFIID, which is necessary for RNA polymerase II-driven transcription. TFIID consists of TBP and TBP-associated factors (TAFs), which help select promoters and regulate transcription.

TFIID

Binds to the core promoter, recognizing sequences such as the TATA box, Initiator (Inr), or downstream promoter elements.

TFIIB

Acts as a molecular bridge, connecting TFIID (bound to the TATA box) to Pol II to form the preinitiation complex (PIC). Binds to TFIID to optimize possible interaction of RNA pol with TFs and DNA. The DNA is bent and con be recognized by RNA pol due to TFIIB.

TFIIE

Facilitates the assembly of TFIIH, which unwinds DNA and mobilizes the transcription machinery. It is essential for the regulation of transcription initiation, ensuring proper interactions between RNA polymerase II and other factors.

TFIIF

Prevents random binding of RNA pol to non-specific DNA regions. It stabilizes the association of RNA polymerase II with the transcription complex and recruits TFIIE and TFIIH for transcription initiation.

Transcription Initiation

Using the energy from the hydrolysis of ATP, the polymerase “closes” around the unwinded DNA to initiate transcription. Transcription starts when RNA pol detaches from all TF.

Elongation Factors

Allow RNA pol II to move through DNA that is packaged into nucleosomes.

Gene

A sequence of nucleotides on one DNA strand that is transcribed into an RNA molecule.

Transcribing a Gene

To “rewrite” the sequence of DNA nucleotides in one gene using RNA language. This process involves synthesizing a complementary RNA strand from the DNA template, which ultimately leads to the production of RNA molecules that can be translated into proteins.

Promoter (Definition II)

A sequence of nucleotides that precedes the gene (at the gene 5’ side) and that is specific to the strand that carries the gene. RNA pol cannot recognize which strand is the gene but the promoter can help RNA pol identify the strand that needs to transcribed.

mRNA Processing

Preparing the synthesized RNA for translation

Challenges for mRNA processing

mRNA needs to leave the nucleus including only the doing sequence of the protein. mRNA needs to be modified for translation to begin through capping, polyadenylation, and splicing.

Phosphorylation of RNA polymerase

Brings negative charges in the RNApol tail, changes conformation of the protein, attracts other protein. Phosphorylation of RNA pol allows RNA processing proteins to assemble on its tail.

mRNA Capping

After RNA modifying molecules modify the RNA pol, mRNA needs to be modified. This is done through 5’ capping, which facilitates mRNA transport to the cytosol and also correct translation.

RNA Splicing

Removal of introns in the middle of the RNA sequence (only done in Eukaryotic cells). The introns form into a lariat structure (loop) as they are spliced/removed from sequence.

Spliceosomes

How snRNP (small nuclear ribonucleoproteins) tag the beginning and end of introns for removal. Ex. of spliceosomes: U6, U1, U2

Polyadenylation

After RNA is modified and capped, a poly-A tail is added to the 3’ end of mRNA, which enhances stability and facilitates export from the nucleus. It also aids in translation initiation and protects mRNA from degradation.

mRNA Transport

Conjugation of prepared mRNA to a protein complex, the Exon Junction Complex (EJC), to pass the Nuclear Pore Complex.

Protein Translation

Genetic information turned into proteins. This process involves decoding mRNA by ribosomes using tRNA molecules to assemble amino acids in the correct sequence, forming polypeptides that fold into functional proteins.

Ribosomes

Protein Factories - molecular machines found in all living cells. They synthesize proteins by translating messenger RNA (mRNA) sequences into polypeptide chains. Structures composed of RNA (rRNA) and proteins.

Genetic Information → Proteins

Sequences of 3 nucleotides in the mRNA molecules (codon) encode for amino acids. Each arrangement of 3 nucleotides “encodes” for one amino acid.

tRNA molecules

The anticodons and the 3’ end; Anticodons bind to complementary codons in mRNA. Each tRNA is specific for the amino acid it is carrying as it is transferred from the nucleus to cytosol.

tRNA synthase/aminoacyl-tRNA synthetase

A protein that “host” in its structure one amino acid and on tRNA based on its anticodon. Aminoacyl tRNA synthetase is specific for each amino acid to be added. Anticodon binds to codon allowing for an amino acid transfer.

Wobbling

One tRNA can read different codons causing an ability of mismatching.

Structure of a Ribosome

rRNA is the core, forming the A, P, and E sites, and is surrounded by proteins. Ribosomes have a large subunit (tRNA bound) and a small subunit (mRNA bound). A site: aminoacyl-tRNA, P site: peptidyl tRNA, E site: exit site

Translation Initiation

Translation starts when the tRNA carrying the amino acid methionine makes interaction with the first available corresponding codon on the mRNA. AUG is a start codon

Reading Frames

Ways in which the sequence of nucleotides in mRNA can be divided into consecutive, non-overlapping triplets. There are three possible reading frames for a given nucleotide sequence, and the correct frame determines how the sequence is interpreted during translation.

Growth of the Polypeptide Chain

A polypeptide chain is transferred from the P-site tRNA to the new amino acid on the A site t-RNA. This process involves the formation of a peptide bond catalyzed by the ribosome, elongating the chain by one amino acid.

Post Polypeptide Chain Growth

After the chain grows, the ribosome translocates (one subunit at a time) to create a new A site for an incoming tRNA.

Translation End

Translation stops when a stop codons appears. Not recognized by tRNA so it cannot be translated.

Release Factor

Changes activity of the peptidyl transferase: one molecule of H2O is added, releasing the amino acid chain from the tRNA.

Post Translational Modifcations

Occurs after folding and suffers the fate of a newly synthesized protein. Folding, binding, covalent modification, noncovalent modification, etc. Phosphorylation can also occur. Acetylation: the addition of an acetyl group to a protein, often regulating its function and location can also occur.

Phosphorylation

A mechanism to regulate a protein’s activity. Ex. for substrate: RNA pol → TFIIH is the kinase. A process where a phosphate group is added to a protein, often affecting its function and activity.

Protein Degradation

The process by which proteins are broken down into smaller peptides or amino acids, often involving proteolytic enzymes. This is essential for regulating protein levels and recycling cellular components.

Proteasomes

Structures within cells responsible for degrading unwanted or damaged proteins, particularly those tagged with ubiquitin. They play a crucial role in maintaining protein homeostasis. Ubiquitin is a small protein that marks proteins for degradation by proteasomes. Protein degradation is a mechanism to control the amount of a protein in a cell.

Proteasome Barrel

Regulatory Particle (RP): Responsible for substrate’s recognition; Core Particle (CP): responsible for protein degradation, contains protease active sites

Degradation of mRNA

If we degrade mRNA, there won’t be translation of relative gene any longer. This process helps regulate gene expression by preventing the synthesis of proteins corresponding to the degraded mRNA.

Regulation of Gene Expression

A way to “control” how much or how little of a selected gene is transcribed into mRNA, to therefore control how much or how little of a protein the cell will be able to produce. It involves various mechanisms such as transcription factors, enhancers, silencers, and epigenetic modifications that influence the availability of mRNA for translation.

Micro RNA (miRNA)

miRNA halts translation (regulates gene expression), Recognition of selective, RNA sequences leading to mRNA degradation.

Regulation of Gene Expression via Transcription

To eventually reduce or increase transcription, and therefore translation, of selected genes. This is important during development, and to determine a cell’s shape and function. All mechanisms from transcription, mRNA, and protein degradation are used to control protein amounts in a cell.

Specialized Cells

Different from one another but in any organism, cells share the very same DNA. DNA may not be accessed in all the same way.

Transcription Regulators

Proteins that make the region of the DNA within a gene of interest available (or not) for transcription, facilitating the formation of the complex of general transcription factors with TATA box. Controls accessibility of specific genes, controlling a cell’s shape and function. Allow the formation of a mediator complex that controls accessibility to TATA box. TRs move to the nucleus to control the expression of selected genes.

The Mediator

Specific proteins of the medicator will selectively interact wither either HATS or HDACS. This regulates chromatin architecture and accessibility to TATA box, once the transcription regulator bind to DNA element, and the mediator forms. Can turn off the transcription of genes through recruitment of proteins that do not allow DNA decondensation.

Gurdon Experiment Outcomes

Genes (DNA content) is the same in all the cells, but proteins are expressed differently, depending on the cell type.

Transcription Regulators (II)

Specialized proteins that regulate the transcription of specific genes/proteins. Specific for selected genes, cell specific (to regulate expression of other proteins necessary for development and function), and are important during development in which cells differentiate into specific cell types. Control transcription of specific genes once in the nucleus.

Transcriptional Control

An orchestra of factors to ultimately change DNA, enabling transcription. Initiator site for attachment of RNA pol. Regulatory DNA sequences turn gens on and off → Element/Operator. TRs bind to regulatory DNA sequences.

Gene Activation

Occurs at a distance; Mediator complex of proteins: binds to both transcriptional complex and activator protein, effect of enhancers favor transcription while repressors inhibit transcription; Transcription regulators binds to dedicated element.

Organization of DNA into Chromatin

The process of packaging DNA into a structure called chromatin, which compacts the DNA to fit within the cell nucleus and regulates gene expression by controlling access to genetic material.

Histone Tails

Regulatory elements of histone proteins that can undergo various modifications, influencing the structure of chromatin and affecting gene expression.

Transcription Initiation

Transcription regulators and chromatin remodling. The process where RNA polymerase binds to the promoter region of a gene, marking the beginning of transcription. This phase involves various transcription factors and the unwinding of DNA to allow RNA synthesis.

Chromatin Remodeling

The dynamic process by which the structure of chromatin is altered, allowing access to DNA for transcription, replication, and repair by repositioning or restructuring nucleosomes.

Transcription and Element

When TRs bind to their element, the formation of a mediator occurs. This complex of proteins interact with the specific TR but will bind to chromatin modifying enzymes.

BDNF Transcription

Brain Derived Neurotrophic Factor is a protein and is essential for neuronal integrity. Can be inhibited through condensation and is a form of regulation for expression. Reduced in Huntington’s disease as more REST goes into nucleus, blocking transcription.

Cell Differentiation

The process by which a less specialized cell becomes a more specialized cell type, involving changes in gene expression and cell function. A factor of gene expression.

Stem Cells

Cells that have the ability to differentiate into various specialized cell types and have the potential for self-renewal. Ex. totipotent (embryo and palcenta), pluripotent (embryo any type), multipotent (mesoderm, endoderm, ectoderm), organ specific (organ repair)

Combinatorial Control

How stem cells become differentiated. Only a few TRs are used in the early stages to control expression, including inducing transcription of TRs.

Cell Memory

A positive feedback loop. A case in which transcription factors can activate their own transcription in absence of a stimulus, remembers things!

iPCS cells

Induced pluripotent stem cells (like the ones of the blastocyst) from differentiated cells; they can self-renew and differentiate into various cell types. Used to study brain function, disease, senescence, DNA modifications

Identification of Specific TRs

Re-programming differentiated cells into stem cells with appropriate TRs

Embryonic Stem Cell Identity

Specific factors can maintain ES cell identity (Oct3/4 Sox2 klf4); Embryonic Stem Cells are pluripotent and can differentiate into any cell type.

iPSCs to Organoids

Self-organized cultures of cells that can resemble a specific tissue or organ, derived from iPSCs. Models human diseases.

Organoids

3D structures that mimic organ physiology and function. Generated by reprogramming one patient’s skin cells (fibroblasts) into iPSC, that are later differentiated into a dedicated set of cells and are developed from human tissues. Transcription regulators help differentiate specific cells in organoids development.

Brain Organoids

Cells are organized in layers, resembling cellular organization in the brain.

miRNA

Downregulation of miRNA can cause neurodegenerative diseases. Biogenesis of miRNA: problems and disease! Any step that is defective will compromise the formation and transport of miRNA, leading to malfunctioning cellular processes and contributing to various disorders. miRNA can treat diseases and are transported through nanoparticles.

Mouse Blastocysts and iPSCs

Injection of selected iPSCs into mouse blastocysts leads to embryo formation and development

iPSCs for cell therapy

Autologous (self to self), allogenic (self to others)

Evolution of Genes and Genome

Role of coding and non-coding sequences in evolution, gene duplication, and horizontal gene transfer.

Rearrangement of DNA sequences

Affects evolution and phenotype; can lead to gene diversity and new functions.

DNA Changes

Alterations in DNA sequences that can impact genetic information, leading to variations in traits and functions. Modifications in any region of a DNA molecule can change how the information stored in DNA is used. Consequences on evolution and disease.

Germ Line Cells (Gamete)

Information in germ line cells will be passed on to the next generations and will define somatic cells. Modifications in somatic cells will not be passed on. Every time that gametes carry “Variations” in the DNA, the genome is “changed” and genes may be changed too. Formation of new genes underlies development and evolution.

New Genes

New genes formed single nucleotide changes in the DNA and rearrangements of DNA sequences.

Point Mutations

A single nucleotide change in either any one exon or intron of a gene, or a regulatory region. They can lead to changes in the amino acid sequence of the resulting protein, potentially altering its function and creating a new protein. Could be toxic (Causing disease) or enign (protective from disease). Ex. lactose in tolerance → point mutation in DNA of LACTASE

Clustered Regularly Interspaced Palindromic Sequences (CRISPR)

Gene editing → a technology that allows for precise modifications to DNA sequences, enabling targeted changes in genes for various applications. A specific DNA sequence found ONLY in the DNA of bacteria cells. Designed to integrate genetic material from viruses into the DNA of bacteria to kill them. Bacteria uses the material to kill the viruses, allowing CRISPR to be an immune system for bacteria.

Virus Invasion in Prokaryotic Cells

The process by which viruses enter and replicate within prokaryotic cells, often leading to the destruction of the host cell. This mechanism is a critical aspect of bacterial evolution and can trigger immune responses such as CRISPR. Viral DNA is transcribed, proteins are made, copied by DNA pol, and viruses are multiplied. A cell infected with a virus releases thousands copies of that virus and the infection propagates,

Bacteria Cell Protection

Bacteria cells exploit the genetic information stored in viral DNA to break down viral DNA itself. Bacteria incorporate sequences of viral DNA within their own DNA, using it later to fight that specific virus.

CAS Proteins

CRISPR-associated proteins that play a crucial role in the CRISPR immune system of bacteria. These proteins are involved in the recognition and cutting of foreign viral DNA, facilitating the editing and protection functions of CRISPR. Transcribed together with the DNA region within CRISPR.

CAS Transcription

The result of such transcription is a set of mRNAS that will be translated into CAS proteins, and a seperate set of different RNA molecules (called cRNA), and used to target foreign viral genetic material for destruction.

CAS Proteins

Designed to break phosphodiester bonds and unwrap DNA. Breaks phosphodiester bonds, helicase/single strand nuclease activity, RNA guided DNA endonuclease

Mature CRISPR RNA

To target nucleic acids, the viral DNA! With CRISPR, no RISC, but a set of transcribed and translated proteins, named CAS proteins, that have a similar function as RISC proteins. With CRISPR, no microRNA involved. Instead, a GUIDE, RNA will be transcribed from the viral genetic information.

Human Genome

Does not contain CRISPR regions or encode for CAS proteins.

Transfections

One plasmid that can encode for both CAS and CRNA and on there with two plasmids.

Membranes

Membranes contain and delimit cells. To separate the extracellular environment from the intracellular one, membranes need to be water impermeable. Intracellular membranes need to be water impermeable.

Intracellular Membranes

Each organelle has a specific function regulated by different proteins and therefore they need membranes to separate them from the rest of the cell. Membranes can carry out specific functions.

Plasma membrane

A biological membrane that separates and protects the interior of a cell from its external environment, regulating the movement of substances in and out of the cell. Consists of lipids and proteins.