The interactome of KRAB zinc finger proteins reveals the evolutionary history of their functional diversification.

Krüppel-associated box (KRAB)-containing zinc finger proteins (KZFPs) are encoded in the hundreds by the genomes of higher vertebrates, and many act with the heterochromatin-inducing KAP1 as repressors of transposable elements (TEs) during early embryogenesis. Yet, their widespread expression in adult tissues and enrichment at other genetic loci indicate additional roles. Here, we characterized the protein interactome of 101 of the ~350 human KZFPs. Consistent with their targeting of TEs, most KZFPs conserved up to placental mammals essentially recruit KAP1 and associated effectors. In contrast, a subset of more ancient KZFPs rather interacts with factors related to functions such as genome architecture or RNA processing. Nevertheless, KZFPs from coelacanth, our most distant KZFP-encoding relative, bind the cognate KAP1. These results support a hypothetical model whereby KZFPs first emerged as TE-controlling repressors, were continuously renewed by turnover of their hosts' TE loads, and occasionally produced derivatives that escaped this evolutionary flushing by development and exaptation of novel functions.

KAP1 is an antiparallel dimer with a functional asymmetry.

KAP1 (KRAB domain-associated protein 1) plays a fundamental role in regulating gene expression in mammalian cells by recruiting different transcription factors and altering the chromatin state. In doing so, KAP1 acts both as a platform for macromolecular interactions and as an E3 small ubiquitin modifier ligase. This work sheds light on the overall organization of the full-length protein combining solution scattering data, integrative modeling, and single-molecule experiments. We show that KAP1 is an elongated antiparallel dimer with an asymmetry at the C-terminal domains. This conformation is consistent with the finding that the Really Interesting New Gene (RING) domain contributes to KAP1 auto-SUMOylation. Importantly, this intrinsic asymmetry has key functional implications for the KAP1 network of interactions, as the heterochromatin protein 1 (HP1) occupies only one of the two putative HP1 binding sites on the KAP1 dimer, resulting in an unexpected stoichiometry, even in the context of chromatin fibers.

KRAB zinc-finger proteins contribute to the evolution of gene regulatory networks.

The human genome encodes some 350 Krüppel-associated box (KRAB) domain-containing zinc-finger proteins (KZFPs), the products of a rapidly evolving gene family that has been traced back to early tetrapods. The function of most KZFPs is unknown, but a few have been demonstrated to repress transposable elements in embryonic stem (ES) cells by recruiting the transcriptional regulator TRIM28 and associated mediators of histone H3 Lys9 trimethylation (H3K9me3)-dependent heterochromatin formation and DNA methylation. Depletion of TRIM28 in human or mouse ES cells triggers the upregulation of a broad range of transposable elements, and recent data based on a few specific examples have pointed to an arms race between hosts and transposable elements as an important driver of KZFP gene selection. Here, to obtain a global view of this phenomenon, we combined phylogenetic and genomic studies to investigate the evolutionary emergence of KZFP genes in vertebrates and to identify their targets in the human genome. First, we unexpectedly reassigned the root of the family to a common ancestor of coelacanths and tetrapods. Second, although we confirmed that the majority of KZFPs bind transposable elements and pinpoint cases of ongoing co-evolution, we found that most of their transposable element targets have lost all transposition potential. Third, by examining the interplay between human KZFPs and other transcriptional modulators, we obtained evidence that KZFPs exploit evolutionarily conserved fragments of transposable elements as regulatory platforms long after the arms race against these genetic invaders has ended. Together, our results demonstrate that KZFPs partner with transposable elements to build a largely species-restricted layer of epigenetic regulation.