Question

On discovery of a new ATP dependent chromatin remodeling complex, made up of 10 proteins, a scientist wants to determine which of these 10 proteins of the complex has catalytic activity of ATP hydrolysis. What domains should the scientist look for in each of the proteins to identify the catalytic protein?

Answer 1

Chromatin provides both a means to accommodate a large amount of genetic material in a small space and a means to package the same genetic material in different chromatin states. Transitions between chromatin states are enabled by chromatin-remodeling ATPases, which catalyze a diverse range of structural transformations. Biochemical evidence over the last two decades suggests that chromatin-remodeling activities may have emerged by adaptation of ancient DNA translocases to respond to specific features of chromatin. Here, we discuss such evidence and also relate mechanistic insights to our understanding of how chromatin-remodeling enzymes enable different in vivo processes.

The packaging of eukaryotic DNA into chromatin provides a means to partition the genome into transcriptionally active and transcriptionally repressed regions. Different patterns of partitioning allow diverse transcriptional programs to arise from a single genetic blueprint. The establishment of specific chromatin states during the course of development as well as their maintenance through the disruptive events of transcription, DNA replication, and DNA repair require rapid rearrangements of chromatin structure. ATP-dependent chromatin-remodeling enzymes provide a means of generating such changes in chromatin structure.

ATP dependent chromatin remodeling is brought about by the factors called remodelers. Remodelers are DNA dependent motors that utilize energy derived from ATP hydrolysis to non-covalently alter this structure . These enzymes are member of a diverse group of proteins named (SWI/SNF) after the archetypal S. Cerevisiae Snf2 proteins; the Snf2 family. Multiple members of this protein family are present in the sequenced genomes of eukaryotes, of which the chromatin remodeling enzymes form distinct sub groupings . The crystal structure of catalytic domains of the two Snf2 related proteins highlight structural similarities with the RecA domain found in the range of helicasess . Snf2 proteins use the energy of ATP hydrolysis to alter the histone DNA interaction. However, unlike bona-fide helicases, the action of chromatin remodeling enzymes are not generally associated with separation of DNA strands.

Remodelers can in vitro mediate (a) nucleosome sliding, in which the position of nucleosome on DNA changes, (b) the creation of a remodeled state, in which DNA becomes more accessible but histones remain bound, (c) complete dissociation of histone and DNA, or (d) histone replacement with variant histones (for a detailed discussion see below). At the same time, ATP dependent remodelers work in conjunction with histone chaperones and histone modifying enzymes.Currently, four different classes of ATP-dependent remodeling complexes can be recognized: SWI/SNF, ISWI, Mi-2, and Ino80. Each class is defined by the presence of a distinct ATPase [10].

SWI/SNF group

Historically, it was the discovery of yeast SWI/SNF complexes in the mid-1980s initiated spurt in studies of chromatin remodeling. First chromatin remodeling complex was purified from yeast. It is product of five SWI and SNF gene (SWI1, SWI2/SNF2, SWI3, SNF5 and SNF6) were found to be constituents of a 2 MDa complex named SWI/SNF complex. Later on affinity-purified complex contained, in addition to SWI1, SWI2/SNF2, SWI3, SNF5 and SNF6, five more then-unknown proteins with molecular weights of 78, 68, 50, 47 & 25 kDa [28].

All prototype SWI/SNF-type complexes studied so far contain a minimal structural and functional core composed of four evolutionarily-conserved subunits: homolog of yeast proteins SWI2/SNF2 (the ATPase, major catalytic subunit), SNF5, SWI3 and SWP73. The complex has an ATPase activity that is stimulated by DNA (~30 fold) or by nucleosomes (~40 fold) . Functional characterizations of the complex revealed that it could stimulate binding of GAL4 (and GAL4 derivatives) to nucleosomal binding sites in presence of ATP. In a mutated complex, wherein the SWI2/SNF2-NTP binding motif is rendered non-functional by a point mutation (K798→A), fails to stimulate activator binding to nucleosomes. This suggests that the ATPase activity of SWI2/SNF2 is essential for the SWI/SNF function, but is not needed for structural assembly of the complex. The complex was found (i) to bind DNA in a sequence-non-specific manner with preference for four-way junction DNA, (ii) to interact with DNA through the minor groove, and (iii) to induce positive supercoiling in relaxed plasmids in the presence of ATP . The complex was however redundant when multiple transcription factors bind to nucleosomes in vitro . Reportedly, the yeast SWI/SNF complex (i) disrupted a nucleosome in the presence of ATP, and (ii), persistently remodeled a specific GAL4-binding site-containing nucleosome along an array of nucleosomes in presence of ATP and GAL4, and (iii) evicted histones from activator-interactive nucleosomes in the presence of an activator . In addition, the complex was found to slide nucleosome along a longer DNA fragment . The available data indicate that the subunits have specific roles in determining the range of targets and biological functions of the complexes.

SWI/SNF group of remodelers can be further subdivided into two distinct highly conserved subclasses. One subfamily comprises yeast SWI/SNF, Drosophila BAP (Brm associated proteins) and mammalian BAF complex; whereas the second family includes yeast RSC, Drosophila PBAP, and mammalian PBAF complexes . Chromatin remodeling activity although well-established across the animal phyla has also been reported in plant.