Parts of the nucleus o Nucleoplasm region of nucleus that is not nucleolus o Chromatin fibers o Nucleolus o Outer nuclear membrane continuous with ER making perinuclear space continuous with ER lumen o Inner nuclear membrane o Perinuclear space 20 40 nm wide o Nuclear pore complexes 3000 to 4000 direct contact between cytoplasm and nucleoplasm channel formed with 30 different proteins called nucleoporins octagonal symmetry 8 spokes extend from center granule 120 nm wide o Nuclear lamina Interphase chromosomes occupy their own territory To be able to see if this is true in normal cells Fluorescence in situ hybridization FISH Each chromosome occupies a distinct territory and is arranged in an organized fashion o The positiosn of these territories do not seem to be fixed vary from cell to cell of same organism and change during a cell s life cycle reflecting changes in gene activity in different chromosomes Space in and around these domains are interchromosomal territory Nuclei are divided into discrete functional domains that play an important role in regulating gene expression and in replication Heterochromatin in periphery Nucleolus has fibrils and granules o Fibrils are DNA being transcribed into rRNA and granules are rRNA being packaged with protein to form ribosomal subunits Nucleolus contain NOR nucleolus organizer region o Stretch of DNA carrying multiple copies of rRNA genes o Humans have 10 NORs per nucleus Instead of 10 nucleolus there are loops of chromatin derived from ten separate chromosomes Size of nucleolus correlated with level of activity During mitosis nucleolus disappears reappears after cell division is done Small non membrane enclosed nuclear structures o Cajal bodies also machines o Gemini of cajal bodies Involved in maturation and processing of snoRNA snRNA o Speckles interchromatin granule clusters rich in RNA and proteins necessary for splicing of messenger RNA precursors into mature messenger RNAs o Promyelocytic leukemia bodies DNA repair o Replication is accurate because of DNA polymerase 3 having proofreading exonuclease activity o Mismatch repair Mut S o Repair occurs after damage by radiation such as thymine thyamine bonds o If mutation repair enzymes are damaged mutation rate increases cancer correlation Point mutation single base change o Missense changes amino acid sequence o Silent does not change amino acid sequence o Frameshift mutation single addition or deletion alters meaning of subsequent codons o Nonsense becomes a stop codon Mutations o Beneficial increase fitness o Neutral no effect on fitness o Deleterious lower fitness Chromosomal level mutations o Inversion segment becomes flipped and rejoined o Translocation segment attached to a different chromosome o Deletion segment of chromosome is lost o Duplication additional copies of a segment are present Base excision repair o Corrects single damaged bases in DNA o Deaminated bases are removed bby DNA glycosylases which recognize deaminated base and remove it by cleaving bond between base and sugar it is attached to o Repair endonuclease detects depurinating breaks phosphodiester bond Nucleotide excision repair o Proteins recognize major distortions in DNA double helix and recruit and enzyme called NER endonuclease that makes two cuts in the DNA backbone on either side of distortion then dnna helicase bings to the stretch of DNA between the nicks and unwinds it freeing it from the rest of DNA gap filled with dna polymerase sealed by dna ligase Nonhomologous end joining sets of proteins bind to end of dna fragments and join them cant prevent loss of nucleotides not sure if correct fragments Homologous recombination uses non broken chromosome as template Only small particles 12 5 nm are allowed through w o npc Nuclear proteins are synthesized by ribosomes in the cytosol and contain a zip code a molecular address tag that marks them for transport through the npc NLS o In nucleoplasmin its in the tail o 8 to 30 amino acids in length positively charged basic contains proline and lysine and arginine importing into mitochondria o transit signal sequence at n terminal o cleaved off by transit peptidase o contain positively charged amino acids secondary sequence important hydrophobic and hydrophic parts when seuqne ce is coiled into alpha helix most charge on one side and hydrophobic on the other Tom translocase in outer mito membrane Tim translocase in inner mito membrane Chloroplast toc and tic Where do the proteins go o Export Ribosomes become attached to RER and growing polypeptide chains are then transferred across or in the case of integral membrane proteins inserted into the ER membrane as synthesis proceeds cotranslational process First you have an er signal sequence that directs the ribosomemRNA polypeptide complex to surface of RER then as polypeptide elongates it crosses er membrane and enters lumen Er signal sequences 15 30 amino acids positively charged N terminal central hydrophobic region and a polar region adjoining the site where cleavage from the mature protein will take place place SRP binds to signal sequence and blocks further translation SRP binds to receptor allowing ribosome to become attached to its receptor GTP binds to SRP and SRP receptor unlocking translation and causing transfer of the signal sequence to the pore protein whose central channel opens as the signal sequence is inserted GTP hydrolyzed accompanied by SRP As polypeptide elongates it passes into ER lumen and signal peptidase cleaves signal sequence which is quickly degraded After polypeptide sequence is completed the final polypeptide is released into ER lumen and translocon channel is closed and ribosome diassociates o BiP attaches to hydrophobic regions so they don t freak out o Protein disulfide isomerase is catalyzes formation of disulfide bridges starts to work even before polypeptide is done figuring out most stable arrangements o Cytosol or mitochondria chloroplasts peroxisomes and nuclear interior Remain free in cytosol polypeptides released and use special targeting signals