Cathepsin L Stabilizes the Histone Modification Landscape on the Y Chromosome and Pericentromeric Heterochromatin

Abstract

Posttranslational histone modifications and histone variants form a unique epigenetic landscape on mammalian chromosomes where the principal epigenetic heterochromatin markers, trimethylated histone H3(K9) and the histone H2A.Z, are inversely localized in relation to each other. Trimethylated H3(K9) marks pericentromeric constitutive heterochromatin and the male Y chromosome, while H2A.Z is dramatically reduced at these chromosomal locations. Inactivation of a lysosomal and nuclear protease, cathepsin L, causes a global redistribution of epigenetic markers. In cathepsin L knockout cells, the levels of trimethylated H3(K9) decrease dramatically, concomitant with its relocation away from heterochromatin, and H2A.Z becomes enriched at pericentromeric heterochromatin and the Y chromosome. This change is also associated with global relocation of heterochromatin protein HP1 and histone H3 methyltransferase Suv39h1 away from constitutive heterochromatin; however, it does not affect DNA methylation or chromosome segregation, phenotypes commonly associated with impaired histone H3(K9) methylation. Therefore, the key constitutive heterochromatin determinants can dynamically redistribute depending on physiological context but still maintain the essential function(s) of chromosomes. Thus, our data show that cathepsin L stabilizes epigenetic heterochromatin markers on pericentromeric heterochromatin and the Y chromosome through a novel mechanism that does not involve DNA methylation or affect heterochromatin structure and operates on both somatic and sex chromosomes.In eukaryotic cells, pericentromeric chromosomal regions do not completely decondense in the interphase and form heterochromatin, a more condensed and transcriptionally repressed type of chromatin morphologically distinct from decondensed and transcriptionally active euchromatin (17,21,55). Previous molecular and genetic studies had established a set of epigenetic mechanisms such as a special set of histone modifications also referred to as "histone code" (24) and DNA methylation (26) that demarcate silent heterochromatin and transcriptionally active euchromatin, separating one from the other. A key regulatory mechanism that controls heterochromatin formation is the positive feedback loop by which histone H3 trimethylated at lysine 9 (H3me3K9) recruits heterochromatin protein 1 (HP1) through direct interaction with the HP1 chromodomain (2, 27, 30). HP1, in turn, recruits histone H3(K9) methyltransferase Suv39h (24, 54) to methylate adjacent H3(K9). In addition to H3me3K9, other factors can contribute to the stable association of HP1 with chromatin (10, 59). The presence of HP1 at chromosomal loci is sufficient to induce chromatin condensation and gene repression (64). The highest concentration of HP1 and H3me3K9 is found at simple repeats in the vicinity of centromeric regions forming the largest blocks of heterochromatin, also known as pericentromeric constitutive heterochromatin. In addition, H3me3K9 is localized on chromatin clusters associated with certain ty...

Pericentric heterochromatin state during the cell cycle controls the histone variant composition of centromeres

“…We generated two Flp-In TM -3T3 cell lines stably expressing either H2A or H2A.Z tagged with the SNAP epitope. Consistent with previous observations (Bulynko et al, 2006;Dryhurst et al, 2009;Greaves et al, 2007;Rangasamy et al, 2004), both variants showed a similar nuclear distribution and neither accumulated at the PHC at steady-state levels in our model system (supplementary material Fig. S1A,B).…”

“…Therefore, we wanted to determine whether the perturbations in new H2A.Z deposition observed in cells with altered PHC translated into alterations in the steady-state distribution of total H2A.Z. By antibody staining in cells with an unaltered PHC state, both H2A variants displayed similar localization, with even distribution at euchromatin and heterochromatin, consistent with previous observations (Bulynko et al, 2006;Greaves et al, 2007;Rangasamy et al, 2004) (Fig. 5A,B).…”

Split top: A maternal cathepsin B that regulates dorsoventral patterning and morphogenesis

“…A null mutant of ctsb in the mouse is homozygous viable but exhibits significantly increased amyloid plaque formation in a mouse model for Alzheimer's disease. In addition, cathepsins have been found in the nucleus and can proteolytically cleave histones to modulate chromatin and regulate ES cell differentiation (Bulynko et al, 2006;Duncan et al, 2008), as well as regulate the CUX1 transcription factor (Ceru et al, 2010;Goulet et al, 2004). Thus, Ctsba could modulate, through proteolytic cleavage, the activity of transcription factors, chromatin or other factors acting in dorsoventral patterning or morphogenesis.…”

Out-of-frame start codons prevent translation of truncated nucleo-cytosolic cathepsin L in vivo

“…An increasing number of functionally important cytosolic or nuclear proteins are assigned as substrates for the lysosomal protease cathepsin L 7,22,29,30 . However, the biogenesis of bona fide cytosolic or nuclear cathepsin L is not well understood.…”

“…It has been reported to cleave the CDP/Cux transcription factor thereby driving G1/S transition of the cell cycle and by cleavage of 53BP1 cathepsin L might contribute to genomic instability in triple-negative breast cancer cells 7,27,28 . Nuclear cathepsin L seems to influence histone modifications, since epigenetic marks on the Y chromosome are reorganized in cathepsin L-deficient mouse fibroblasts 29 . Furthermore, recent reports demonstrated cleavage of the N-terminal tail of histone H3 by cathepsin L during embryonic stem (ES) cell differentiation 30,31 .…”