In bacterial chromosomes, the position of a gene relative to the single origin 
of replication generally reflects its replication timing, how often it is 
expressed, and consequently, its rate of evolution. However, because some 
archaeal genomes contain multiple origins of replication, bias in gene dosage 
caused by delayed replication should be minimized and hence the substitution 
rate of genes should associate less with chromosome position. To test this 
hypothesis, six archaeal genomes from the genus Sulfolobus containing three 
origins of replication were selected, conserved orthologs were identified, and 
the evolutionary rates (dN and dS) of these orthologs were quantified. Ortholog 
families were grouped by their consensus position and designated by their 
proximity to one of the three origins (O1, O2, O3). Conserved orthologs were 
concentrated near the origins and most variation in genome content occurred 
distant from the origins. Linear regressions of both synonymous and 
nonsynonymous substitution rates on distance from replication origins were 
significantly positive, the rates being greatest in the region furthest from any 
of the origins and slowest among genes near the origins. Genes near O1 also 
evolved faster than those near O2 and O3, which suggest that this origin may 
fire later in the cell cycle. Increased evolutionary rates and gene 
dispensability are strongly associated with reduced gene expression caused in 
part by reduced gene dosage during the cell cycle. Therefore, in this genus of 
Archaea as well as in many Bacteria, evolutionary rates and variation in genome 
content associate with replication timing.