Background The role of chromosomal arrangements in adaptation is supported by the repeatable clinal variation in inversion frequencies across continents in colonizing species such as Drosophila subobscura. However, there is a lack of knowledge on the genetic variation in genes within inversions, possibly targets of climatic selection, across a geographic latitudinal gradient. In the present study we analysed four candidate loci for thermal adaptation, located close to the breakpoints, in two chromosomal arrangements of the sex (A) chromosome of Drosophila subobscura with different thermal preferences. Individual chromosomes with A2 (the inverted arrangement considered warm adapted) or AST (the standard ancestral arrangement considered cold adapted) were sequenced across four European localities at varying latitudes, up to ~ 2500 Kms apart. Results Importantly, we found very low differentiation for each specific arrangement across populations as well as no clinal patterns of genomic variation. This suggests wide gene exchange along the cline. Differentiation between the sex chromosome arrangements was significant in the two more proximal regions relative to the AST orientation but not in the distal ones, independently of their location inside or outside the inversion. This can be possibly due to variation in the levels of gene flux and/or selection acting in these regions. Conclusions Gene flow appears to have homogenized the genetic content within-arrangement at a wide geographical scale, despite the expected diverse selective pressures in the specific natural environments of the different populations sampled. It is thus likely that the inversion frequency clines in this species are being maintained by local adaptation in face of gene flow. The differences between arrangements at non-coding regions might be associated with the previously observed differential gene expression in different thermal regimes. Higher resolution genomic scans for individual chromosomal arrangements performed over a large environmental gradient are needed to find the targets of selection and further elucidate the adaptive mechanisms maintaining chromosomal inversion polymorphisms.