Chicken histone H5 is an H1-like linker histone that is expressed only in nucleated erythrocytes. The histone H5 promoter has binding sites for Sp1 (a high affinity site) and UPE-binding protein, while the 3' erythroid-specific enhancer has binding sites for Sp1 (one moderate and three weak affinity), GATA-1, and NF1. In this study we investigated whether trans-acting factors that bind to the chicken histone H5 promoter or enhancer are associated with adult chicken immature and mature erythrocyte nuclear matrices. We show that NF1, but not Sp1, GATA-1, or UPE-binding protein, is associated with the internal nuclear matrices of these erythroid cells. Further, we found that a subset of the NF1 family of proteins is bound to the mature erythrocyte nuclear matrix. These results suggest that in chicken erythrocytes NF1 may mediate an interaction between the histone H5 enhancer and the erythroid internal nuclear matrix. NF1 was also present in the internal nuclear matrices of chicken liver and trout liver. The observations of this study provide evidence that NF1 may have a role in a variety of cell types in targeting specific DNA sequences to the nuclear matrix.