Isochores

particularly bacteria, plants, invertebrates;

little variation among vertebrates

a lot of heterogeneity within their genomes

long stretches (100s of kb) of DNA with uniform %GC

• could span >10Mb

High %GC = heavy isochores (H)

Low %GC = light isochores (L)

L1 (<37% GC),

L2 (37-42%),

H1 (42-47%),

H2(47-52%),

H3(>52%)

L1 and L2 are often grouped together as a single L isochore

only in warm-blooded vertebrates (mammals, birds), not in cold-

blooded vertebrates (fish).

selectionist:

• GC-pairing is stronger than AT-pairing

• may stabilize DNA at higher temperatures

• hypothesis is supported by the observation that heavy isochores are found in warm-blooded vertebrates. Furthermore, heavy isochores are gene-rich

mutationist:

• pool of available nucleotides changes over replication

• There is more GC available early inreplication, so mutations will be biased towards G or C.

• Over time, regions of the genome that replicate early become GC-rich.

• In general, GC-rich regions have beenobserved to replicate early.

• By doing a sliding window analysis of the human genome DNA sequence and defining isochores as segments >300 kb with distinct %GC and low heterogeneity

• 41% of genome covered

• coding regions!

• codings regions > introns > 5’flanking regions > 3’ flanking regions

• Certain codons are “preferred” and are used much more frequently than “unpreferred” codons

• e.g. Leucine

  • 6 different codons, CTG, CTA, CTC, CTT, TTG, TTA

  • expect codon to be used about 1/6

  • CTG instead used 90%

• preferred codons correspond to the most abundant tRNA in each species

  • selection favors the use of codons that increase the level of gene expression

ENC

• effective number of codons, the average number of codons that are used to encode the 20 amino acids

• minimum is 20 (one codon per a.a.) the maximum is 61 (all codons except the 3 stop codons).

• Low ENC = high codon bias.

• ENC can be applied to any species without prior knowledge of expression or codon usage.

Fop

• frequency of optimal codons, the frequency with which the “optimal” codon is used for each amino acid

• Optimal codons = those used with the highest frequency in highly expressed genes.

• High Fop = high codon bias.

• species-specific and requires that optimal codons are known. For this, one must have many gene sequences and expression information.

• higher in highly-expressed genes

• higher in short genes than in long genes

• higher in female-expressed than in male-expressed genes

• natural selection favors the use of optimal codons to make translation faster and more accurate

• codon bias used as a way to regulate gene expression post transcriptionally

evidence:

• highly expressed genes have higher codon bias

• conserved protein motifs have higher bais than other protein regions -> selection for accuracy of translation

Replacement of optimal leucine codons with non-optimal codons leads to a lower level of

ADH protein in vivo and reduces ethanol tolerance in adult flies

(Wa-F = wild-type, optimal leucine codons

1 leu = 1 luecine codon changed from optimal to non-optimal

6 leu = 6 leucine codons changed from optimal to non-optimal

10 leu = 10 luecine codons changed from optimal to non-optimal

In a comparison of ADH protein concentration, it was found that:

Wa-F > 1 leu > 6 leu > 10 leu)

The Finding: Replacing sub-optimal leucine codons with optimal ones increases ADH enzymatic activity in larvae, but decreases it in adults.

The Reason: It demonstrates an evolutionary trade-off. Codon usage is not optimized for a single stage but balanced across the life cycle, though codon bias remains highest in larval-expressed genes because larvae face the highest environmental ethanol stress

• bias in mutation could lead to codon bias

1. The Mutation Bias: Spontaneous chemical mutations constantly push the genome to become rich in A and T.

2. The Mismatch Repair: This mutation pressure creates base-pair errors. When the cell fixes them, the biased mismatch repair machinery chemically favors fixing them back into G-C pairs.

3. The Result: This repair bias overrides the AT-mutation bias, packing synonymous third-codon positions with G and C (especially in high-recombination zones).

4. The Human Context: Because human populations are small, selection is too weak to choose codons (Ns≤1). Thus, what looks like "preferred" GC codons is simply a neutral byproduct of this biased cell repair mechanism.

weak

Ns=1 , N is population size & s is selection coefficient

-> often difficult to distinguish between selective & neutral expkanations for codon bias

yes

• small genomes& large population sizes (bacteria) = selection

• humans & other vertebrates = mutational biases explain more