The Nucleus: Terra Incognita Flashcards
Lecture 21
Describe the structure of the nuclear envelope.
Nuclear envelope is comprised of the outer and inner nuclear membranes and is contiguous with the ER.
Parts of the envelope (both membranes and/or just the inner membrane) invaginate (some form membrane-bounded compartments within nucleus), forming the nucleoplasmic reticulum.
What is the nucleoplasmic reticulum?
parts of the nuclear envelope that have invaginated; function is unknown (could be for calcium storage, lipid synthesis, regulation of gene expression and repair)
What are the functions of the nuclear envelope?
- Compartmentalization
- Site of transport
- Structural integrity
- Regulation of gene expression
- Platform for signaling
What are lamins? What do they do?
protein filaments that form a network in the nucleus called the nuclear lamina, which upholds the structural integrity of the nuclear envelope
What is the nuclear lamina?
thin, dense meshwork of fibers that lines the inner surface of the inner nuclear membrane and confers mechanical strength to the nucleus; problems have been linked to premature aging
What would happen if the nuclear lamina were mutated or nonexistent?
The nuclear envelope would be structurally unstable.
What is chromatin?
proteins associated with DNA, forms fibers 10-30 nm in diameter that are dispersed throughout the nucleus
What is a nucleosome?
repeated unit of DNA wrapped around a histone (complex of proteins)
How thick are chromatin fibers in the nucleosome?
10-30 nm thick
What is a chromatin compartment?
when chromatin is packaged together into a territory
What structure do chromosomes take when cells are at rest?
chromosomes decondense and fill the space in the nucleus, arranged in compartments
Describe the experimental evidence for a territorial organization of interphase chromosomes.
2 different potential models for how chromosomes are arranged: mixed like spaghetti vs. separated into territories with no intermixing
Strike the nuclei with an X-ray beam to inflict damage on DNA and see what pattern the damage takes by seeing where the radioactive nucleotide is incorporated via EM radiography.
Rather than radioactivity being randomly distributed, radioactivity was at one-two chromosomes, supporting the territory-organization model
During the X-ray beam laser experiment, what would the evidence look like if the interphase chromosomes were mixed? Was this model true?
Chromosomal damage would be randomly distributed to many different chromosomes. Model was incorrect.
During the X-ray beam laser experiment, what would the evidence look like if the interphase chromosomes were separated into territories? Was this model true?
Chromosomal damage would be on one (maybe two) chromosomes. Model was correct.
Describe fluorescence in situ hybridization (FISH).
Technique: use a small, fluorescently labeled, nucleic acid probe to find out where other nucleic acids are in the cell. Complementary DNA molecule base pairs with given DNA sequence somewhere in the nucleus. Did this with each chromosome, could show where each chromosome was located; noticed each occupied distinct regions in the nucleus.
Which chromatin is not transcriptionally active?
heterochromatin
What are the differences in transcriptional activity between hetero and euchromatin?
Heterochromatin is not active. Euchromatin is active
Which kind of chromatin is transcriptionally active?
euchromatin
What technique provides the best evidence that chromosomes are separated into territories? Why?
Fluorescence in situ hybridization.
Chromosome painting allows each chromosome to be located in vivo.
In contrast, the X-ray laser experiment results could have been coincidental
What are the differences in electron microscopy staining in heterochromatin vs. euchromatin?
Dark regions are heterochromatin and light regions are euchromatin.
What are the different levels of compaction between heterochromatin and euchromatin?
Heterochromatin is compacted. Euchromatin is not compacted.
Can a single chromosome be both transcriptionally active and inactive?
Yes. Some areas can be active while other areas are not active.
Why isn’t euchromatin compacted?
Transcriptionally active DNA can’t be compacted. To be transcribed, it must lose its histones.
When DNA is not wrapped in histones, what structure does it take?
Naked DNA often congregates in certain areas of a chromosomal territory called transcriptional hubs.
What are transcriptional hubs?
when DNA is naked, preparing to be or in the middle of being transcribed, it often goes to certain places (neighborhoods) within the chromosomal territory
What happens to the location of genes while their transcribed?
The positions of genes within their transcriptional hubs changes when transcribed.
What moves chromatin into different areas within the transcriptional hub during transcription?
Diffusion
What is the position effect on gene expression?
difference in transcription between certain areas within a chromosomal territory; moving between areas also changes transcriptional activity
Why isn’t chromatin free to move around in the nucleus?
Because it’s attached to the lamina, which provides structural integrity to the nuclear envelope.
What are the drivers of nuclear organization?
Polymer self-interactions
Chromatin binding proteins
Homotypic clustering (eu and heterochromatin found in different areas in the cell)
What are the constraints of nuclear organization?
Nuclear envelope (chromatin attached to lamina)
Nuclear bodies
What are nuclear bodies?
an ordered region within the nucleus in spaces with a lot of transcriptional activity, where proteins move onto the DNA; a constraint to nuclear organization
How many different kinds of nuclear bodies are there?
over a dozen
Name 3 nuclear bodies.
Nucleolus
Cajal body
Speckles
What happens at the nucleolus?
rRNA production/ribosome subunit assembly
What is a cajal body?
nuclear body - assembly of complexes required for mRNA processing
What are speckles?
nuclear bodies - storage of complexes required for mRNA processing
In the nucleus, does structure determine function or does function determine structure?
Function determines structure.
What technique provides evidence that the position of genes changes when transcribed?
Fluorescent in situ hybridization
What are histones?
small proteins with positively charged amino acids (lysine and arginine) that encourages the binding of DNA
How does histone acetylation impact packing? What about histone methylation?
Histone methylation leads to tighter packing. Acetylation leads to looser packing, promoting chromatin decondensation.
When there is a problem with the nuclear lamina, what kind of effects does it have on humans?
premature aging (progeria)
How does the nuclear envelope help organize chromatin?
Part of it are bound to the inner nuclear envelope at sites associated with the nuclear pores; these regions are heterochromatin
