HMG EXAM 2 Combined Set

Compare and contrast the structure of mitochondrial and nuclear DNA 

• Mitochondrial DNA: circular, 37 genes, heavy and light strands. Organized in nucleoids. The heavy strand is guanine rich and the light strand is cytosine rich. Has a D-loop where replication occurs twice on heavy strand. Has a small triplex segment. RNA product is multigenic. Maternal origin. 66% protein coding. 

• Only one copy per nucleus. Linear/double helix formation, Much larger with many more genes. Genes don’t really overlap. Either Paternal or maternal origin. HIghly conserved sequence. 1.1% protein coding. 

Describe the general process of mitochondrial replication (it may be helpful to draw this as you talk it out) (3 ish steps)

1) Start at CRD, H strand replicates until it reaches L strand
2) L strand replicates in opposite direction
3) Meanwhile, H strand continues past the start point, which results in the CRD loop being replicated twice

What part of the mitochondrial genome is replicated twice?

CRD loop

What nuclear DNA mechanism is MtDNA replication intermediate repair mechanism similar to?

Okazaki fragment resolution

Name two proteins involved in MtDNA repair and nuclear DNA repair

FEN1, DNA2

Name a few similarities between MtDNA and nuclear DNA

tRNA and rRNA genes, wobble, GC rich

What is the endosymbiotic theory?

That mitochondria are descended fromj basteria that fused with nucleated cells.

Name at least three pieces of evidence that suggest the endosymbiotic theory is true?

Each have their own genome, mtDNA do not organize into nucleosomes, inhibitors of bacterial translation block mitochondrial translation, but not nuclear translation. New mitochondria are formed by binary fission. Comparisons of rRNA gene sequences suggest mitochondrial genomes derive from a common ancestor of α-proteobacteria.

There are ? tRNA genes, ? rRNA genes, and ? protein coding genes in MtDNA

There are 22 tRNA genes, 2 rRNA genes, and 13 protein coding genes in MtDNA

A pedigree shows an affected father passing the disease to affected children. Should you suspect a mitochondrial disease? 

NO, only mothers can pass on mitochondrial diseases. 

A mother has a diagnosed mitochondrial disease. She has identical twin daughters; however, one of them has more severe symptoms, and the other has mild symptoms. Additionally, their symptoms manifest in different organ systems, but are thought to be associated with the same disease. Explain how this is possible. 

Heteroplasmy and replicative segregation. 
Heteroplasmy means that the clinical severity of the disease is directly rrelated to the proportion of mutated MtDNA 

What type of symptoms might someone with MERRF (Myoclonic epilepsy and ragged red fiber disease) have 

Brain, heart, skeletal muscles (type 1 and 2) and skin tissue symptoms. Lack of energy

What is a mitochondrial disease associated with irreversible vision lost

Leber hereditary optic neuropathy (LHON)

Draw a gene with two exons and label the following : enhancer, promotor, 5’ end, 3’ end, 5’ UTR, 3’ UTR, splice donor site, splice acceptor site, exon, intron, start codon, stop codon, TATAA box, CAAT box, GC box. 

Actually try drawing it without looking at your notes. This will likely be a labeling question. 

Describe the difference between an enhancer and a promotor. 

A promotor is directly upstream of the transcription start site, usually pretty close. Enhancers can be kinda anywhere that’s not an exon. It can be in the UTRs or it can even be in the intron. 

What’s the difference between a focused and dispersed TSS 

Focused is only one promoter, with one point of transcription initiation. A dispersed TSS can have multiple promotors. 

T/F A mutation that prevents the expression of a primary pioneer transcription factor for a gene that codes for a protein involved in regulating neuronal development would likely be less deleterious than a mutation in one of the gene’s many regular transcription factors. 

Likely to be false. A mutation in a primary transcription factor means that the DNA will not be open to allow other TFs to bind to and activate that region 

A multifactorial error occurs that causes 3D chromatin formation to not form topologically associating domains. How would you expect that to affect gene expression?

Can result in inappropriate activation or repression as a lack of regulation.

Are all control elements for a specific gene active in all types of tissues?

No. For example a gene could have one enhancer activating it moderately in one tissue type, another enhancer acting it mildly in another, or a suppressor and other regulating elements silencing the gene in a third tissue type.

BRE function (b-recognition elements)

Helps recognize where TATAA box or TSS is

GC box function

GC rich seq that acts as a binding site for transcription activators like SP1 and acts like an enhancer element to help recruit RNA Pol II to promoters that dont have TATA box

TATA box function

Found in 15% of all genes, AT rich sequence that helps recruit TBP and RNA Pol II to TSS 

Name the trigger and purpose of replicative senescence

Telomere erosion, prevents cancerous proliferation

Name the trigger and purpose of oncogene-induced senescence

Oncogene overexpression, prevents early formation of tumors

Name the trigger and purpose of stress-induced senescence

Mitochondrial dysfunction or inflammation, and rapid response to cellular stress and prevents damaged cells from dividing

Name the trigger and purpose of therapy induced senescence 

Trigger is chemotherapy and radiation, and the purpose is to prevent further tumor growth 

What is the rate of proliferation in labile cells?

Continuous

What is the rate of proliferation in permanent cells?

No proliferation

Describe the G1/S checkpoint generally

1) Is there enough room to grow, are conditions favorable?

During the G1/S checkpoint, Cyclins ? and ? need to be expressed at [high/low] levels to move through the cycle

cyclins E and A need to be expressed at high levels to move through the cycle

During the S-Phase checkpoint, what inhibits cyclin E/A and therefore stops the replication origins from firing

CDC25A

What is the cell checking during the G2/M checkpoint? Describe generally. 

Is DNA replication complete? Is it correct? 

During the metaphase checkpoint of mitosis, what is the cell checking? Describing generally.

Have all chromosomes arrived and aligned at the metaphase plate?

During the G2/M checkpoint, ATM/ATR signals CHK1/2 to [inhibit/promote] Cdc25B/C. Increased Cdc25B/C signaling is needed to [inhibit/promote] the forward progression of the cell cycle into mitosis. 

During the G2/M checkpoint, ATM/ATR signals CHK1/2 to inhibit Cdc25B/C. Increased Cdc25B/C signaling is needed to promote the forward progression of the cell cycle into mitosis. 

What specifically does the cell check for at the metaphase checkpoint? 

Tension on the kinetochores. 

How is the intrinsic pathway of apoptosis triggered? 

Chemical or radiation damage 

If we are discussing capsases activating capsases, what are we probably talking about?

Apoptosis

What is the trigger for the extrinsic pathway of apoptosis?

Another cell signals for the cell to undergo apoptosis by putting external proteins on the cell to trigger downstream apoptotic cellular messaging. 

Cytochrome C leaving the mitochondria is associated with the [intrinsic/extrinsic] pathway of ?

Intrinsic pathway of apoptosis 

cfDNA comes from ?

Cells that have undergone apoptosis

T/F Does a single cell differentiate into a different cell in one cell lifetime. Ex. can a blood cell differentiate into a skin cell?

No. And a good study question is to try and explain why the answer is no.

Totipotent cells can differentiate into ? Does this include extra-embryonic membranes

All possible cells and extra embryonic membranes

Pluripotent cells can differentiate into ? Does this include extra-embryonic membranes

All possible cells of the embryo, but not extra embryonic structures 

Multipotent cells can give rise to

Multiple lineages, but not all body cells

Oligopotent cells can give rise to

A few types of differentiated cell

Unipotent cells can give rise to?

One type of differentiated cell only.

OEQ: What is an advantage/purpose of cell differentiation?

Introduces complexity to allow cells to become specialized

What is the relationship between cell cycle checkpoints and cancer

When cell cycle checkpoints start failing it can be one of the ways that cancer develops.

alternatively, if the cell is leaning towards cancer (due to other things going wrong) mutations that promote inhibition of checkpoints will be advantageous to the cancer becoming more cancerous

What is the difference between senescence and quiescence? 

Quiescence is temporary and reversible, and may be due to lack of nutrition and growth factors whereas senescence is due to aging and serious DNA damage 

Compare and contrast between Meiosis and Mitosis

Meiosis: specialized germ line cells, haploid sperm and egg products, only one round of replication per two cell divisions, prophase can take decades to complete, pairing of maternal and paternal homologs, recombination occurs normally, genetically different daughter cells

Mitosis: Occurs in all tissues, creates diploid somatic cells, normally one round of replication per one cell division, short prophase (30 min), rare and abnormal to have recombination, daughter cells are genetically identical 

When is maximum condensation for mitosis and meiosis?

Metaphase for both. 

What is the purpose of the spindle checkoint?

Ensures that chromosomes are properly attached to the spindle before cell moves onto anaphase

What is the centrosome made of

Centrioles, centrosome matrix, microtubules, and gamma tubulin ring complex

What is the kinetochore?

Protein complex surrounding the centromere for kinetochore microtubules to attach to

What are two  spindle checkpoint inhibitors? (stops separin from activating)

Mad2 and BubR1

What is the complex that activates securin which activates separin?

APC (Anaphase promoting complex)

What are the five steps of synapsis

1. Pairing

2. Crossing over

3. Partial separation (DHJ) 

4. Chiasmata

5. Segregation of homolog 

Describe purpose of recombination

Diversity. ((Equity inclusion and justice) /j)

Key features of germ cells

Pluripotent, can be reduced to haploid after meiosis. In testes, every round produces 4 spermatozoa. In ovaries, division is asymmetric.

List chromosomal abnormalities resulting from errors of recombination and nondisjunction

I was gonna come back and answer this but i feel like y’all probably know this.

What is heterochromatin

Closed/more condensed, hypoacetylated

What is euchromatin

Open, hyperacetylated

T/F Nucleosomes cannot shift in position, they are completely stationary

False, they can move around relative to DNA to either expose or hide DNA segments

Where are the methyl groups added in DNA methylation?

To the cytosine bases, especially in CpG islands by DNA methyltransferase

Name two similarities and two differences between chromatin and DNA methylation.

Similarities: Methyltransferase enzyme is used to add methyl groups. Methylation is used as a method of regulation to turn genes on or off. 
Differences: DNA methylation occurs on cytosine whereas chromatin methylation occurs on the lysine (usually) of histone tails. DNA methylation does not alter the heterochromatin/euchromatin state of DNA whereas chromatin methylation can. Chromatin methylation also has three different versions of methylation (tri mono and di) 

T/F When methylated, CpG islands are GC rich regions of DNA in promoters that provide binding sites for transcription factors

False, this only happens when unmethylated

Testing shows that a gene has chromatin that is heavily methylated. Can you assume that the gene is activated or repressed?

No. Chromatin methylation can both activate or repress gene expression, depending on the specific amino acid residue modified .

T/F Testing shows that a gene has chromatin that is heavily acetylated. Can you assume that the gene is activated or repressed?

Yes you can, and it is activated (euchromatin)

T/F Euchromatin promotes gene expression.

True. Euchromatin = open

eRNA role

transcriptional regulation

rRNA role

Protein synthesis and export (ribosomes)

snRNA role

Part of the spliceosome  

snoRNA role

Chemical modification of rRNA and snRNA (like methylation), transcribed from introns

asRNA role

(antisense) binds to mRNA to prevent translation

lncRNA role

Regulates gene expression in the nucleus and in the cytoplasm.

miRNA role 

Perfect complementation to target leads to degradation, mostly perfect complementation leads to binding and sequestering. 

piRNA role

transposon control + can affect chromatin

siRNA role

arises from dsRNA for transposon control as well as RNAi-based gene silencing. Silences the mRNA by inducing a cut via AGO2

shRNA

artificial RNA used to trigger RNA interference

tRNA role

binds to codon to create proteins 

Organize the following RNA from smallest to largest: tRNA, miRNA, rRNA

miRNA (21-22nt), tRNA (70-80nt), rRNA (120-500nt)

Name three similarities and two differences between DNA and RNA polymerases

Similarities: Use DNA template, both add nucleotides to the 3’ end and move in a 5’-3’ direction. Both use watson-crick base pairing rules.

Differences: DNA Pol is used in DNA replication whereas RNA pol is used for transcription. RNA Pol adds ribonucleotides rather than deoxyribonucleotides. RNA Pol doesn’t need primers.

Name the three processing steps for mRNA

5’ cap (7-methyl guanine), splicing via snRNPs, cleavage and polyadenylation

The most common RNA found in the nucleus

pre mRNA transcripts

what elements of mRNA protects it from degradation 

5’ cap protects from 5-3’ exonucleases, 3’ polyA tail adds stability with length, and the 3’UTR has elements that regulate decay (some microRNAs can bind to complementary sequences in the 3’ UTR) 

What are the three options for mRNA decay

1) Deadenylation independent ARE mediated decay (cap is removed)
2) Deadenylation dependent ARE mediated decay (Poly A tail is degraded—> exosome for degradation)
3) Duplex formation with a second RNA molecule that leads to either [cleavage + exonuclease degradation] OR [signaling via Ago that directs nucleases to destroy the RNA]

What are the five steps of microarrays?

1. Isolate mRNA

2. Make labeled cDNA

3. Hybridize to microarray

4. Scan fluorescence

5. Analyze data

What is the purpose of a microarray?

To measure expression levels, which can be visualized by the amount of fluorescence produced

What IS RNA editing?

Altering the nucleotide sequence through insertion deletion or base substitution, possibly turning adenosine to inosine or cytosine to uridine.

What is the purpose of RNA editing

Increases protein diversity as well regulate stability and splicing. ie. if RNA is edited at a specific site it can create or destroy a splice site which affects alternative splicing. 

Compare and contrast the function and mechanism of siRNA and miRNA 

siRNA (bind and cut) complexes with AGO2
miRNA (bind and degrade or bind deadenylate/sequester) complexes with argonaut

3 Functions of the 5’ UTR

1) contains hairpin structures that can interact with proteins to control translated regions
2) contains and establishes upstream open reading frame (uORF) which modifies levels of translation of coding product of mRNA
3) contains internal ribosome entry site (IRES) where the ribosome loads the mRNA onto the kozak consensus sequence

Describe the three functions of the 3’ UTR

1) Contains hairpin loops, like the 5’UTR, which can interact with proteins and control translated regions 
2) Contains the polyadenylation site which adds stability
3) Interacts with antisense RNA to influence subcellular localization 

what is the IRE

Iron response elements, which can upregulate or downregulate elements depending on iron binding 

What are protein signal sequences

Sequences of amino acids that dictate where protein is sorted. 

A protein signal sequence contains an extended block of hydrophobic amino acids, where might the protein be moved?

Imported into the ER

A protein signal sequence contains many lysines and an arginine, where might the protein be moved?

Imported into the nucleus

What are three irreversible post translational modifications?

Deamidation, eliminylation, proteolysis

What are six reversible post translational modifications 

Methylation, acetylation, phosphorylation, hydroxylation, ubiquitylation, glycosylation  (MAP HUG) 

What amino acids are phosphorylated? Is it reversible?

Tyr, Ser, Thr. Yes