DNA elements of chromosomes

Origins of replication

-          where DNA replication starts

-          although DNA replication is fast (50nt/sec in humans), eukaryotic genomes are huge, so need lots of oris

-          mammalian – 10,000 oris

o       DNA replicating out from one ori until it hits the replication fork from the adjacent oris is a replicon

-          Some oris used every replication, some used in only some replications

o       Early embryonic cells tend to do mitosis very rapidly, so need to replicate DNA rapidly, so lots of origins firing

-          Some oris fire early during replication, others later

-          Can isolate oris by putting genomic DNA sequences on a plasmid that lacks an ori – if plasmid is maintained and passed on to both daughter cells in mitosis it must have an ori

-          Yeast oris relatively simple

-          Other eukaryotic are hard to figure out

Telomeres and telomerase

-          problem of telomere replication

o       leading strand can replicate to end, lagging strand has RNA primer at end

o       this is chewed away, leaving a single strand

§         single strand gets chewed away, therefore chromosome gets shorter each replication

o       so why don’t chromosomes get shorter with every replication?

-          Telomere structure

o       Elizabeth Blackburn - telomere sequences are lots (kb) of repetitive sequence

§         Humans (and mammals, birds, reptiles, plants) it is TTAGGG

·         Suggests evolved a long time ago

§         Tetrahymena it is TTGGGG

-          telomerase

o       this enzyme maintains telomeres

§         Carol Greider and Blackburn

o       it is a ribonucleoprotein (like snRNPs or ribosomes)

§         has proteins and also a piece of RNA that is an integral part of it

o       Works in the way outlined in Fig. 13.10

§         This is a DNA polymerase that carries its own template

§         Adds nt 5’ to 3’ just like any DNAP

§         Template is the piece of RNA

§         Adds on to overhanging 3’ end left by replication

o       Must be some mechanism to regulate telomere length – so that when they start to get too short telomerase adds some sequence

-          is this the cure for cancer?

o       fully differentiated tissues have telomerase turned off (so have a limited life span)

o       cancer cells from these tissues turn telomerase back on

-          is this the key to aging?

o       if put telomerase into fully differentiated cells in tissue culture, they live forever

Centromeres

-          DNA sequence that gets hooked up to the spindle during mitosis

-          Always lots of repetitive sequence, noncoding, known as satellite sequence

-          Short sequences of 5-300bp, repeated many times

o       A million bp

-          Seems to be packaged in a different way than typical chromatin – extremely condensed

o       During metaphase can actually see as a constriction in a chromosome (Fig. 13.12b)

-          Lots of proteins bind to this region

o       Cohesin – binds and maintains pairing of sister chromatids

o       The kinetochore – protein/centromere structure that hooks chromosomes to the spindle in mitosis

Interesting problems….

4, 5a, 7, 8, 10a, 12, 13, 15, 17, 21