Boundaries of eliminated heterochromatin of Tetrahymena are positioned by the DNA-binding protein Ltl1

• Published in: Nucleic acids research 2019-08, Vol.47 (14), p.7348-7362
• Main Authors: Jaspan, Vita N, Taye, Marta E, Carle, Christine M, Chung, Joyce J, Chalker, Douglas L
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Binding Sites - genetics; Cell Line; DNA, Protozoan - genetics; DNA, Protozoan - metabolism; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Gene Expression Regulation; Gene Knockout Techniques; Gene regulation, Chromatin and Epigenetics; Heterochromatin - genetics; Heterochromatin - metabolism; Protein Binding; Protozoan Proteins - genetics; Protozoan Proteins - metabolism; Sequence Homology, Amino Acid; Tetrahymena thermophila - cytology; Tetrahymena thermophila - genetics; Tetrahymena thermophila - metabolism
• ISSN: 0305-1048; 1362-4962
• DOI: 10.1093/nar/gkz504

Abstract During differentiation of the Tetrahymena thermophila somatic nucleus, its germline-derived DNA undergoes extensive reorganization including the removal of ∼50 Mb from thousands of loci called internal eliminated sequences (IESs). IES-associated chromatin is methylated on lysines 9 and 27 of histone H3, marking newly formed heterochromatin for elimination. To ensure that this reorganized genome maintains essential coding and regulatory sequences, the boundaries of IESs must be accurately defined. In this study, we show that the developmentally expressed protein encoded by Lia3-Like 1 (LTL1) (Ttherm_00499370) is necessary to direct the excision boundaries of particular IESs. In ΔLTL1 cells, boundaries of eliminated loci are aberrant and heterogeneous. The IESs regulated by Ltl1 are distinct from those regulated by the guanine-quadruplex binding Lia3 protein. Ltl1 has a general affinity for double stranded DNA (Kd ∼ 350 nM) and binds specifically to a 50 bp A+T rich sequence flanking each side of the D IES (Kd ∼ 43 nM). Together these data reveal that Ltl1 and Lia3 control different subsets of IESs and that their mechanisms for flanking sequence recognition are distinct.