Genome And The Cell Flashcards
Which is the short arm and which is the long arm of the chromosome?
P is short
Q is long
What are the 5 non coding DNA groups?
Non-translated RNA (micro RNA, long non coding RNA)
Chromatin binding sites
Transposons
Promoter and enhancer regions (for binding of transcription factors etc)
Special structural regions (centromeres, telomeres etc)
What is euchromatin?
Actively transcribed area of chromosome
What is heterochromatin?
Transcriptionally inactive part of the chomasome
What is a nucleosome?
DNA wrapped around a histone core. 147bp to be specific.
What are the 2 most common types of DNA variation in human?
Single nucleotide polymorphism and
Copy number variations
What does the P stand for with regard to arms of a chromasome?
Petite. This is how you know it is the smaller arm. The Q arm is just named because it is next in the alphabet.
What are the histone “marks” that impact DNA accessibility for transcription (i.e. euchromatin)?
Methylation, acetylation, phosphorylation, specific amino acid absence/presence
What amino acids within histone are subject to methylation? What affect does this have on accessibility for RNA polymerase to transcribe DNA?
Lysines and arginines in histone can be methylated. Methylation of lysine residues can lead to either increased or reduced DNA accessibility depending on which residue is methylated.
What amino acid residues within histone are subject to acetylation? What affect does this have on accessibility for RNA polymerase to transcribe DNA?
Lysine residues are acetylated by histone acetyltransferases. These, typically, open the chromatin for increased transcription. Histone deacteylases HDACs lead to condensation of chromatin and reduced transcription. HDACs are being targeted by cancer treatments as their overactivation (and so gene silencing) appears to be critical to cancer development.
What amino acide residues with histone are subject to phsophorylation? What affect does this have on accessibility for RNA polymerase to transcribe DNA?
Serine residues can be modified by phosphorylation. Depending on which serine residues is phosphorylated will dictate whether this contributes to euchromatin or heterochromatin formation.
Other than histone modification, what else can epigenetically effect the accessibility of DNA to RNA polymerase?
DNA methylation
What is the effect of DNA methylation?
Transcriptional silencing. This a tightly regulated process.
What do chromatin organising factors do?
Bind to noncoding regions and DNA to coordinate long range looping to bring promotor and enhancer regions together to influence transcription.
How many sub componenets of histone are there?
8 - it’s an octamer. 9 if you count the H1 linker moleculres.
Is this more likley to be euchromatin or heterochromatin?
Euchromatin. Acetylation of histone lysine residues is associated with the open chromatin state able to be transcribed. Selectly positioned lysine methylation, and selctly positioned serine phosphorylation can also contribute to the euchromatin state.
Is this more likely to be euchromatin or heterochromatin?
Heterochomatin. Extensive methylation of histone lysine and arginine residues is consistent with a closed chromatin formation unable to be easily transcribed. Select phosphorylation of serine residues can also contribute to heterochromatin formation.
Are histones positvely or negatively charged?
Positivley. This allows negatively charged DNA to stick to it.
What steps are involved in the production of fucntional miRNAs?
Transribed from DNA to 1) Priamry miRNA.
Cleaved in the nucleus into 2) Pre-miRNA.
3) Exported out of the nucleus via export protein.
4) In the cytoplasm, trimmed by the protein ‘Dicer’.
5) Now miRNA at final length, associates with a protein complex known as RISC (RNA-induced Silencing Complex). RISC + miRNA can now bind to imperfectly matched mRNA in the cytoplasm to impair its translation at the ribosome, or perfectly matached RNA to cleave it.
What happens when the RNA-induced Silencining Complex (RISC) + miRNA imperfectly binds to mRNA in the cytoplasm?
It causes the mRNA to be unable to be translated via ribosomes, so its corresponding gene is silenced.
What happens when RNA-induced Silencing Complexes (RISCs) +miRNA bind perfectly to mRNA?
The RISC cleaves the target mRNA at the binding position, silencing the gene for which the mRNA was a transcript.
What are siRNAs?
Small interefering RNAs. They are sequences that can be injected into cell cytoplasm to by trimmed by Dicer, then interact with RISC, experimentally, thus forming miRNA RISC complexes and gene silencing artificially.
What is long non-coding RNA?
Recently discovered long transcripts of non-coding DNA that alters chromatin function amongs many other things. Best known clinically significant example is XIST - a long non-coding RNA (lncRNA) that inactivates the X chromosome as part of normal X-inactivation in females (complete gene silencing of one X chromosome). XIST is, interestingly, derived from the inactived chromosome but doesn’t cloak its own expression.
What functions does long non-coding RNA have?
Gene activation - by facilitating transciption facotr binding
Gene suppression - by intercepting transcription factors and preventing them from binding to DNA.
Chromatin modification to indirectly impact transcription (image depicts two way this has been shown to occur - through direct aciton on methylases an acetylases, or through stabilisation of complet protein structures that allow chromatin changes).
What are CRISPRs? What is does Cas refer to?
Clustered, regularly interspaced, short palindromic repeats. (CRISPRs)
CRISPR associated genes (Cas)
– the best known is CAS9, which is a nuclease, which brakes DNA of viral or plasmid invaders in prokaryotes.
What is the prokaryote physiological role the CRISPR Cas-9 combination?
In bacteria, CRISPRs are interpaced with sequences derrived from small broken parts of phage or plasmid DNA are inserted into the prokaryotic genome by a Cas1-Cas2 complex. This then allows the bacterial production of small targetting RNA based on the phage/plasmid specific spacers that complex with other Cas (e.g. 3, 9, 10) enzymes and are used to home in on the specific phage/plasmic DNA. The complexed Cas enzyme then destroys the DNA by cleaving it (they are nucleases).
How can the CRISPR-Cas 9 system be used for gene editing?
Artifical guide RNAs can be devloped to target sections of DNA to be cleaved, then complexed with Cas9. Cas9 will cleave the target sequence out of the DNA, in the same way that would to cleave and destroy an invading plasmid or phage.
What can CRISPR-Cas9 be used for?
1) Removal of phathogenic genetic material
2) Addition of genetic material - benefical therapeutically, of for investigation purposes in the lab.
Where are proteins destined for membrane bound status or secretion synthesised?
Rough endoplasmic reticulum and the golgi apparatus
What does smooth endoplasmic reticulum do?
Specifically, it used for the synthesis of steroid hormones and lipoprotines, as well as modification of hydrophobic compounds into water soluble compounds.
Where are proteins bound for the cytosol synthesised?
Free cytosolic ribosomes
What is the role of proteasomes?
They are protein complexes that degrade denatured or other proteins that have been tagged for destruction. The resulting protein products are displayed on MHC, and often can influence intracellular signalling pathwasys.
What is the role of lysosomes?
Lysosomes are intracelular packets of degradtive enzymes that fuse with endocytotosed components (e.g. microbes) to destroy them, but are also the site of intracellular organelle breakdown as part of normal cellular component recycling (autophagy). They contain enzymes that are capable of degrading a very large range of macromolecules.
What is the role of peroxisomes?
Intracellular packets of oxidative enzymes (e.g. catalase, peroxidase) important for the breakdown of very long chain fatty acids. They produce hydrogen peroxide in the process.
What are the key structural proteins intracellularly?
Filamentous actin (microfilaments)
Keratins (intermediate filaments)
Microtubules
What is meant by cell polarity?
The orientation of the cell relative to other structures. Critical for coordinated cell function to complete tasks like form a cell layer. Apical refers to the outer most surface. Basal is the deepest surface. Lateral are the other surfaces. Each surface requires the deployment of different proteins, so maintaining orientaiton/polarity with cytoskeletal structures (microtubules, microfilaments and kertatins) is critical.
What functions are completed within mitochondria?
Oxidative phosphoylation and ATP production
Production of metabolic intermediates
Syntesis of some macromoleculres (e.g. heme)
Contain important sensors for cell damage - the mitochondria can then responsd by initiating apoptosis.
What is the lifespan of a mitochondria?
10 days
What phospholipid heads are typically on the cytosolic surface of the bilayer?
Phosphatidyl-ethanolamine
Phosphadidyl-serine - important for maintaining the intracellular negative charge of the membrane.
What phospholipid bilayer components are typically on the extracellular face?
Glycolipids
Sphingomyelin
Phosphatidyl-choline
What phospholipid bilayer components are seen in the inner and out most layers about equally?
Cholesterols
Phosphatidyl-inositol
—when intracellular and phosphorylated, this forms an important intracellular signalling molecule scaffold. They can also hydrolysed intracellularly by phospholipase C to generate second signals like diacylglycerol and inositol triphosphate.
What happens to cells that have a lot of phosphatidylserine displayed extracellularly?
Phosphatidylserine is displayed almost entirely intracellularly in healthy cells. When displayed extracellularly, this is a signal of cell stress and attracts other sells to destroy them in a controlled way (e.g. phagocytosis).
What do glycosylphosphatidylinositols do?
GPIs link proteins to phospholipid bilayers extracellularly.
Which part of the phospholipid bilayer is negatively charged?
The inside.
What is caveolin?
It’s a protein concentrated with lipid rafts that helps make calveolae along the outer surface along a phospholipid bilayer capable of endocytosing extracellular fluid, some membrane bound proteins, and some receptor bound moleculels. GPI-linked moleculres, cyclic adenosine monophosphate (cAMP) binding proteins, src-family kinases and the folate receptor are also required for calveolae formation. They can subsequently fuse with endosomes and/or recycle back to the membrane. This process referred to as caveolae (little caves) mediated endocytosis.
What is clatharin? What does it do?
It creates a coating of the internal surface of the phospholipid bilayed deep ot receptors on the external surface involved in receptor-mediated endocytosis. The clathorin allows for internalisation of the external components and endosome formation. Clatharin-coated pits require a protein called dynamin to release the new vesicle into the cytosol.
In receptor mediated endocytosis, how is the ligand released from the receptor in teh endosome to enable it to be used by the cell?
Endosomes become progressively more acidic which disturbs the attraction between the protein-based receptor and its ligand. Depending on the ligand, their are different passive and active ways of then moving the ligand out of the endosome. The receptors can then be recycled back to the membrane. Alternatively, the endosome can fuse with lysosomes which smash apart the receptor to get the ligand out.
What is the role of receptor-mediated endocytosis in down-regulation of receptors on cell surface?
For cell surface receptors that bind protein for endocytosis, the resultant endosome can be directed to fuse with lysosome which will destroy both the receptor and the ligand. The net outcome of this, unless there are counterregulatory pathways that coexist, is downregulation of the receptor pathway due to receptor destruction.
Do actin microfilamnets extend at their negative or positive end?
G-actin monomers noncovalently polymerase into F-actin (the polymer known as microfilament) that then intertwines to forma double stranded helices with a positive and negative end. New subunits are added to the positive end, and removed from the negative end. Sometimes called actin treadmilling.
How do actin filaments move in muscle tissue?
ATP driven hydrolysis of actin by myosin slides the actin filaments relative to one another leading to movement e.g. muscle contraction.
Do intermediate filaments move like microtubules and microfilaments?
No - they form a rigid structure to provide tensile resistence for other things to anchor to, and to provide strength to the cell so it doesn’t break under force.
Do intermediate filaments interact in adjacent cells?
Yes, through structures called desmosomes. They stick together at these points forming a strong superstructure.
Why are intermediate filiments helpful for determining cell line of origin when looking at tumour tissue?
The have specific patterns depending on the cell of origin. These have different names.
E.g. Vimentin: in mesenchymal cells (like fibroblasts and endothelium)
Desmin: in muscle cells - forms the scaffold that actin and myosin sit on.
Neurofilaments: critical fo neuronal axon structure and confer boht stegnth and rigdity.
Glial fibrillary acid protein (GFAP) - is expressed in glial cells specifically.
Cytokeratins - expressed in epithelial cells - 30 subtypes specifically expressed in different tissues (lungs, GI epithelium etc).
Lamins - form the nuclear lamina, define nuclear shape and can regulate transcription.
What is vimentin, and where is it typically found?
It’s a cytoskeletal structure belonging to the group known as a intermediate fibres. It is found in, and can be used to identify, mesenchymal cells, such as endotherlium and fibroblasts.
What is desmin and where is it found?
It’s a cytoskeletal structure belonging to the group known as a intermediate fibres. It is found in, and can be used to identify, muscle tissue. It forms a scaffold for actin (a dynamic cytoskeletal rope) and myosin (an enzyme that manipulates and moves actin).
What are cytokeratins, and where are they found?
They are a cytoskeletal structures belonging to a group known as a intermediate fibres. There are over 30 subtypes of cytokeratins that are found in, and can be used to identify, a range of different tissues including lung and GI tract.
What is laminin and where are they found?
It’s a cytoskeletal structure belonging to the group known as a intermediate fibres. It is responsible for the shape of cell nuclei, and can influence DNA transcription.
What is Glial fibrillary acidic protein?
It’s a cytoskeletal structure belonging to the group known as a intermediate fibres. It is found in glial cells in the central nervous system and can be used to identify these cells.
List the three cytoskeletal protein components in order of size
Smallest
1) actin microfilaments 5-9nm across
2) intermediate filaments 10nm across
3) microtubules 25nm across
Largest
What are microtubules made of?
Proteins called alpha and beta-tubulin that are polymerised together.
Are microtubules active or passive structures?
Extremely active.
What are the characteristics of microtubules?
They are polarised (+/- ends)
The -ve end is embedded in the microtubule organiszing centre and is paired with centrioles.
The +ve end elongates or reced in response to stimuli be the addition or subtraction of tubulin dimers.
What is the role of microtubules in the movement of molecules and organelles inside a cell?
They are mooring lines for molecular motor proteins that use ATP to do work - translocate vescicles, organelles or other molecules around the cell. The most famous of these motor proteins are the kinesins (transport cargo anterograde - away from the nucleus - from the -ve to +ve ) and dyneins (tranpsort cargo retrograde, that is toward the nucleus, from the +ve to the negative end of the microtubule).
What are kinesins? What do they do?
Kinesins are motor protein complexes that associate with microtubules to transport other porteins, moleculres or organelles inside cells. They move typically in the ‘anterograde direction’ - that is from the negative to the positive end of the microtubule they are associated with. There counterpart dyneins moves in the other direction.
What is dynein? What does it do?
Dynein is a motor protein complex that associates with intracellular microtubules to tranport other proteins, organelles or moleculres around the cell. It does this by moving along microtubules in the retrograde (toward the nuclear) direction, from the positive end to the negative end of the microtubule. It’s counterpart kinesinn moves in the other direction.
What is a MOMP?
Mitochondrial outer membrane permeabilisation protein. When the accumulation of pro-apoptotic signals reaches a threshold, and overcomes antiapoptotic signalling, the mitochondria produce lots of MOMP, which allows the escape of cytochrome C amongst other intramitochondrial proteins.
How do notch receptors influence change on cell function?
Notch receptors have and intracellular and extracellular domain joined across the plasma membrane. They bind their counterpart notch ligand on another cell via the extracellular domain. When bound, the extracellular part is snapped off, and this releases the intracellular domain free into the cytosol. It traffics to the nucleus and acts directly as a transcription factor.
How does the Wnt/Frizzled receptor system work?
Receptor specific to wnt receptor extracellular domains becomes bound. Conformational change of wnt leads to association with frizzled, which recruits another intracellular protein called dishevelled, which releives intracellular beta-catenin from a protein complex that usually consituitively inactivates it using ubiquitin tagging. With dishevelled impairing this complex, beta-catenin is then free to traffic to the nucleus and exert transcriptional effects.
What are the primary regulator proteins of the cell cycle?
Cyclins and cyclin dependent kinases (CDK)
Apart from collagen IV, what other proteins are a critical component of basement membrane?
Laminins. These can interact with receptors of cells adjacent to the basement membrane and trigger intracellular signalling cascades. Proteolgycans are also there.
