Individual anti-H1(0)
monoclonal antibodies were screened in an immunolocalization assay to isolate clones able to recognize
H1(0) in a differentiation-dependent manner using a murine
erythroleukemia cell line. Two clones were selected, one recognizing
H1(0) only in differentiating cells (clone 27 antibody), and the other recognizing the
protein constitutively (clone 34 antibody). Both
antibodies recognized a restricted region of the
protein located at the N-terminal part of the globular domain.
Amino acids 24-30, essential for the recognition of the
protein by the clone 27 antibody, are extremely conserved in all known H1(0)-like
proteins from sea urchin to human. Within these residues,
proline 26, responsible for a bend in this region, plays a particularly important role in the
epitope recognition. The region involved in the
protein recognition by clone 34 antibody is larger and encompasses
amino acids 20-30. However,
proline 26 does not play an essential role in the structure of this
epitope. Detailed analysis of the differential recognition of
H1(0) in
chromatin during cell differentiation and proliferation suggests that the modification of
chromatin structure as well as that of
H1(0) conformation can account for this effect. Indeed, in vitro study of H1(0)-four-way junction
DNA interaction showed that the N-terminal tail domain of the
protein can influence the recognition of
H1(0) by these
antibodies when the
protein interacts with
DNA. The two
monoclonal antibodies described here therefore seem to be valuable tools for investigating fine modulations in
chromatin structure and the concomitant changes occurring in the conformation of the
protein.