GEN 2: Defining the Genome I - Chromosomes Flashcards
Observe the learning outcomes of this session
Briefly describe mitosis
Stages:
- interphase
- prophase
- metaphase
- anaphase
- telophase and cytokinesis
- https://www.yourgenome.org/facts/what-is-mitosis
Briefly describe meiosis
Stages:
- interphase
- prophase I
- metaphase I
- anaphase I
- telophase I and cytokinesis
- prophase II
- metaphase II
- anaphase II
- telophase II and cytokinesis
- https://www.yourgenome.org/facts/what-is-meiosis
List some cytogenetic methods used to visualise chromosomes?
- metaphase spreads
- G-banding
- fluorescent in situ hybridisation (FISH)
Why are chromosomes captured during metaphase to produce metaphase spreads?
- during most of the cell cycle, nuclear chromosomes are de-condensed and cannot be observed as distinct entities
- during metaphase, they are condensed and oberservable
- cultured human cells have the cell cycle blocked and halted in metaphase
- they are then spread onto microscopic slides for counting
What are the basic features of metaphase chromosomes?
What is a karyotype?
- the number and appearance of a complete set of chromosomes
What is G-banding?
- the most common method for determining the karyotype of a cell
- uses Geisma dye to stain chromosomes, to produce light and dark bands that reflect structural differences in the chromosome
- allows pairs of chromosomes to be distinguished by their:
- length
- centromere position
- banding pattern
What are cells with a normal chromosome number called?
- euploid
What are cells with an abnormal chromosome number called?
- aneuploid
Describe fluorescent in situ hybridisation (FISH)’s role in visualising chromosomes
- fluorescent signals allow us to visualise the location and determine the intensity
- DAPI stain is used to identify the nucleus
- fluorescently labelled DNA probes can be hybridised to visualise specific chromosomal regions
What kind of chromosomal abnormalities may occur?
- deletion
- duplication
- inversion
- substitution
- translocation
What are some of the parallels between Mendel’s ‘units of inheritance’ and chromosomes, and what did this suggest?
- see diagram
- it suggested that genes are carried by chromosomes
What are the features of X-linked recessive inheritance?
- males affected by X-linked recessive disorders much more frequently
- no male-male transmission
- all daughters of affected males are carriers
- around 50% of female carriers’ sons are affected
- around 50% of carriers’ daughters are carriers
Select the correct statements about meiosis
The chromosome theory of inheritance proposed that Mendel’s factors (genes) are located in chromosomes because (choose the best answer):
Select the correct statements
What are chromosomes made up of?
- chromosomes are made up of chromatin, which is:
- a complex of DNA, protein and RNA
Does the extent of chromatin condensation vary?
- yes, it is highly variable
- interphase chromatin is generally more decondensed than metaphase chromatin, but not uniformly de-condesnsed
What is heterochromatin?
- darkly stained-highly condensed chromatin
What is euchromatin?
- lightly stained-light condensed chromatin
How do you think the level of chromatin condensation impacts gene transcription?
- heterochromatin is highly condensed and usually transcriptionally inactive
- euchromatin is more ‘open’ and able to bind to transcription factors
What are constitutive heterochromatin regions?
- regions of the genome that are heterochromatic at virtually all times and in all cells
- generally centromeric and telomeric regions
What are facultative heterochromatin regions?
- regions of the genome that may form heterochromatin in one cell type and euchromatin in another
- since not all genes need to be transcribed in all cell types at all time
Are ‘house-keeping’ genes heterochromatic or euchromatic?
- they are euchromatic in all cell types
Compare the properties of euchromatin and heterochromatin
- compaction
- cytogenetic appearance
- G-band staining
- presence of genes
- GC/AT content
- replication timing
- repeat sequences
- histone acetylation
- DNA methylation
What are the weight proportions of histone proteins and DNA in chromatin?
- chromatin contains similar weights of histone proteins and DNA
What are nucleosomes formed by?
- nuclear DNA wraps around histone octamers (molecules each of H2A, H2B, H3 and H4)
How many fold can DNA be condensed up to?
- up to 10,000 fold
What controls chromatin condensation?
- the post-translational modification of N-terminal histone tails:
- acetylation
- methylation
- phosphorylation
How do histone acetylation and phosphorylation make DNA more accessible to transcriptional proteins?
- nucleosomes are normally positively charged and attracted to the negatively charged DNA backbone
- histone acetylation and then phosphorylation, respectively, will remove the positive charge and introduce a negative charge
- this facilitates chromatin decondensation, as it weakens the histone tail-DNA interactions
- making DNA more accessible to transcriptional proteins
What does histone methylation do?
- histone methylation increases hydrophobicity
- encoourages chromatin condensation and transcriptional silencing
Apart from histone methylation, what else silences genes?
- DNA methylation: occurs at cytosine residues in CpG dinucleotides
What are some specialised variant histones roles?
- H2A variant, H2AX, forms nucleosomes that are phosphorylated at the site of DNA double-strand breaks
- this marks regions of DNA damage and facilitates chromatin remodelling and DNA repair
Do chromosomes only associate with histones proteins?
- no, they associate with many non-histone proteins
- they contain a similar amount of histone and non-histone porteins
Briefly describe the general roles of the cohesin complex
- hold sister chromatids together
- controls chromatin condensation
Until when does cohesin keep sister chromatids attached?
- cohesin ensures sister chromatids are attached during mitosis, but only until anaphase
What is the cohesin complex made up of?
- a complex of five proteins:
- Smc1 and Smc3: which are two ‘structural maintenance of chromosomes’ (Smc) proteins
- three non-Smc proteins
- arranged in a ring-like structure
What are the two models cohesin may link sister chromatids together?
- Simple Embrace:
- each ring encircles the two sister chromatids entirely - Handcuff:
- each sister encompassed by distinct rings that then interact with each other
Describe the role of cohesin and how it relates to the stages of mitosis
Apart from chromatid cohesion, what is another role of cohesin>
- it plays an important role in chromatin folding/unfolding mechanisms by bringing sequences far apart on a DNA molecule into close proximity to form loops
- this is called the ‘loop extrusion model’
Explain the ‘loop extrusion model
- DNA is pushed through the two rings of extrusion complex (a double cohesin complex)
- the size of the loops is determined by interactions with a transcription factor, CTCF, which binds specific DNA sequence motifs
What does the ‘loop extrusion model help explain?
- helps explain how different regions of the genome can adopt different states of compaction and gene activity in different parts of the nucleus
How many nucleotides do long ncRNA have?
- lncRNA have >200 nucleotides
What is the lncRNA that regulates X-chromosome inactivation?
- Xist
What is the Barr Body?
- as females inherit two X-chromosomes, one of them is randomly inactivated during development
- the inactive X-chromosomes (Xi) is heterochromatic and during interphase, in seen as a highly condensed structure called the Barr body
How is Xist lncRNA related to the Barr body?
- the Barr body is coated with Xist lncRNA, but not the active X-chromosome
- during development, the 17-20kb lncRNA is transcribed from only one of the two X chromosomes
- it acts in cis to promote its heterochromatisation
- Xist only coats and inactivates the X-chromosome it is transcribed from
- Xist binds to proteins that promote methylation and deacylation of histones
Transcriptionally inactive chromatin tends to have:
Observe this diagram of the endosymbiotic theory of mitochondrial evolution
How much of our DNA is found in the mitochondria?
What types of proteins do they code for?
- a relatively small proportion of our DNA is found in the mitochondria
- but it is crucial for our health because it codes for some key proteins in the inner mitochondrial membrane, vital for ATP synthesis via the electron transport chain and oxidative phosphorylation
- e.g. ATP synthase and cytochrome oxidase
How many copies of its chromosome are there in each mitochondrion?
- there are around 2-10 copies of its chromosome in each mitochondrion
What shape is mitochondrial DNA?
- it is circular
- it is not packaged into chromatin
Why does mitochondrial DNA have a faster mutation rate?
What are the implications of this for studies?
- it is replicated less accurately than nuclear DNA
- it has implications for studies of human evolution and ancestry
How many base pairs long is the mitochondrial chromosome?
- 16, 569 base pairs (bp) long
How many genes does the mitochondrial chromosome code for?
What are its functions?
- 37 genes:
- 13 for proteins necessary for ATP synthesis
- 24 for RNA molecules specifically for mitochondrial protein synthesis
Can mitochondria survive with their 37 genes?
- those genes are essential, but not sufficient for mitochondrial functions
- most proteins for oxidative phosphorylation and other mitochondrial processes are encoded by nuclear genes
Observe the mitochondrial chromosome map
Why is the proportion of all human DNA that is mitochondrial DNA almost 10%?
Describe why these are the features in a mitochondrial inheritance pedigree:
- inheritance is only through maternal lines
- affected males do not pass on the genes
- since sperm mitochondria never enter the egg during fertilisation, a father can never pass on a mitochondrial mutation to his offspring
- all affected individual inherit the mutation from their mothers
Compare the nuclear genome and the mitochondrial genome
- genome size (kb)
- gene number
- DNA topology
- gene density
- genetic code used
- chromosome copies
- gene organisation
- inherited from
