S3:

1
S3 Meet the Molecules

• Nuclear genes
• ITS ribosomal internal transcribed spacer
• Glu Polyphenolic adhesive protein
• Mitochondrial gene
• CO3 Cytochrome c oxidase subunit III

2
The molecules

• ITS the internal transcribed spacer of the
  nuclear ribosomal genes
• Ribosomal genes code for rRNA
• Spacer regions are transcribed but then removed
• Region has restriction site polymorphism between
  species

http//fp.bio.utk.edu/mycology/Techniques/mt-what_
dna.htm
3
ITS
http//fp.bio.utk.edu/mycology/Techniques/mt-what_
dna.htm
4
ITS
Multiple copies in genome many targets for PCR
http//fp.bio.utk.edu/mycology/Techniques/mt-what_
dna.htm
5
ITS
PCR product
M. galloprovincialis (959 bp)  
442 156
172 189
    M. trossulus (951 bp)   197
240 156
169 189

• Restriction sites and fragment lengths for the
  Mytilus ITS locus using Hha I restriction enzyme
• ITS is under less selection ? more variable!
• Reference Braby Somero, 2005, Marine Biology

6
ITS
M. galloprovincialis (959 bp)  
442 156
172 189
    M. trossulus (951 bp)   197
240 156
169 189
442
240 197
7
The molecules

• Glu Polyphenolic adhesive protein - nuclear
• Mefp-1 protein
• One of the glues that attaches byssal threads to
  rocks
• Number of repeats (and therefore gene length)
  varies between species

repeat
protein
8
Glu

• Adhesive proteins anchor byssal treads to hard
  surface
• At least five different proteins
• Many variants of each
• Most studied foot protein is Mefp-1
• The Mefp-1 gene codes for the polyphenolic
  adhesive protein

http//biomaterials.bme.northwestern.edu/mussel.as
p
9
Glu
Collagen-like threads with adhesive proteins
http//biomaterials.bme.northwestern.edu/mussel.as
p
10
Glu
DOPA groups (hydroxylated tyrosines) chelate
metal ions on surface
http//biomaterials.bme.northwestern.edu/mussel.as
p
11
Novel uses for mussel adhesives

• Safe, natural glue for surgeries
• Coating for medical implants
• Tooth coating for decay prevention
• Anti-biofouling for boats
• But synthesis is difficult
• Sticky
• Post-translational modification

12
Mefp-1 protein
110 kDa

• Decapeptide repeats contain multiple
• Lys Lysine electrostatic
• Hyp Hydroxyproline sticky
• DOPA hydroxylated tyrosine chelator
• Repeats vary in number with 80/protein

13
Mefp-1 gene
160
2502 bp
474
654

• Glu primers amplify
• 2 products in invader M. galloprovincialis
• 300 and 500 bp (shown above)
• 1 product in native M. trossulus
• 240 bp
• Number of repeats (and therefore gene length)
  varies between species

14
The molecules

• 3. CO3 Cytochrome c oxidase subunit III -
  mitochondrial
• One subunit of a large protein in the electron
  transport chain
• Species level differences are observed after DNA
  sequencing

15
Mitochondria

• Organelles involved in oxidative respiration to
  produce ATP
• Derived from bacterial endosymbionts (about 1.2
  billion years ago)
• Have genome, a circular DNA molecule (as in
  modern bacteria)

http//www.bioeng.auckland.ac.nz/images/database/b
ioinformatics/mitochondria.gif
16
Mitochondrial DNA (mtDNA)

• Most ancestral genes lost or explanted to nucleus
• Remaining genes
• Proteins for respiration
• Ribosomal genes
• Mitochondrial DNA qualities
• A haploid genome
• Non-Mendelian inheritance
• Little or no repair, therefore, evolves 4 x
  faster than nuclear DNA
• Useful for evolutionary studies of closely
  related species

http//users.rcn.com/jkimball.ma.ultranet/BiologyP
ages/E/Endosymbiosis.html
17
Cellular respiration
Complex I Complex II Complex III Complex IV
Subunits NADH Dehydrogenase Cytochrome bc1 Succinate Dehydrogenase Cytochrome c Oxidase
mtDNA 7 1 0 3
nDNA 35 10 4 10
TOTAL 42 11 4 13

18
3D structure of cytochromes
http//dec4.ucdavis.edu/website/Bioenergetics/