We report here the molecular and cytological characterization of two proteins, 
ScoHET1 and ScoHET2 (for Sciara coprophila heterochromatin), which associate to 
constitutive heterochromatin in the dipteran S. coprophila. Both proteins, 
ScoHET1 of 37 kDa and ScoHET2 of 44 kDa, display two chromodomain motifs that 
contain the conserved residues essential for the recognition of methylated 
histone H3 at lysine 9. We raised antibodies to analyze the chromosomal location 
of ScoHET1 and ScoHET2 in somatic and germline cells. In S. coprophila polytene 
chromosomes, both proteins associate to the pericentromeric regions and to the 
heterochromatic subterminal bands of the chromosomes. In germinal nuclei, 
ScoHET1 and ScoHET2 proteins distribute to the heterochromatic regions of the 
regular chromosome complement and are abundantly present along the 
heterochromatic germline-limited "L" chromosomes. We investigated histone 
methylation modifications and found that all heterochromatic regions enriched in 
ScoHET1/ScoHET2 proteins exhibit high levels of di- and tri-methylated histone 
H3 at lysine 9. Taken together, our results support that the association of 
ScoHET1/ScoHET2 to heterochromatin is mediated by histone H3K9 methylation. 
Using 5-methylcytosine antibodies, we proved the cytological detection of DNA 
methylation in S. coprophila. From our observations in L germline chromosomes, 
heterochromatin in S. coprophila is highly enriched in DNA 5-methylcytosine 
residues.