Disruption of NAP1 genes in Arabidopsis thaliana suppresses the fas1 mutant phenotype, enhances genome stability and changes chromatin compaction

Abstract

SUMMARY Histone chaperones mediate the assembly and disassembly of nucleosomes and participate in essentially all DNA‐dependent cellular processes. In Arabidopsis thaliana, loss‐of‐function of FAS1 or FAS2 subunits of the H3‐H4 histone chaperone complex CHROMATIN ASSEMBLY FACTOR 1 (CAF‐1) has a dramatic effect on plant morphology, growth and overall fitness. CAF‐1 dysfunction can lead to altered chromatin compaction, systematic loss of repetitive elements or increased DNA damage, clearly demonstrating its severity. How chromatin composition is maintained without functional CAF‐1 remains elusive. Here we show that disruption of the H2A‐H2B histone chaperone NUCLEOSOME ASSEMBLY PROTEIN 1 (NAP1) suppresses the FAS1 loss‐of‐function phenotype. The quadruple mutant fas1 nap1;1 nap1;2 nap1;3 shows wild‐type growth, decreased sensitivity to genotoxic stress and suppression of telomere and 45S rDNA loss. Chromatin of fas1 nap1;1 nap1;2 nap1;3 plants is less accessible to micrococcal nuclease and the nuclear H3.1 and H3.3 histone pools change compared to fas1. Consistently, association between NAP1 and H3 occurs in the cytoplasm and nucleus in vivo in protoplasts. Altogether we show that NAP1 proteins play an essential role in DNA repair in fas1, which is coupled to nucleosome assembly through modulation of H3 levels in the nucleus.

Histone modifications and chromatin remodelling in plants in response to salt stress

“…FASCIATA1 (FAS1) is one of the subunits of CHROMATIN ASSEMBLY FACTOR 1 (CAF-1). The fas1 mutant possessed a significantly increased H3.3:H3.1 ratio (Kolarova et al, 2020). Although the m123-2 triple mutant alone did not show a significant change in the H3.3:H3.1 ratio, this triple mutation could restore the H3.3:H3.1 ratio to a lower level in the fas1 m123-2 quadruple mutant (Kolarova et al, 2020).…”

Telomerase Interaction Partners–Insight from Plants

“…To explore connections between telomerase and chromatin during cell-cycle progression and/or DNA damage repair we focused on candidate proteins linked to chromatin remodeling. Using yeast two-hybrid (Y2H) assays, we detected protein–protein interactions of two AtTERT constructs, RID and FW3NNLS ( Figure 2 A), with members of the NAP (nucleosome assembly protein) family that function as H2A-H2B histone chaperones [ 25 , 26 , 27 ]. Interestingly, the NAP-related (NRP) family members NRP1 and NRP2 did not interact with AtTERT or POT1a and no interaction was detected in the case of seven other proteins ( Table S1 ).…”

“…To complete our description of new interactions with an overview of previous reports ( Figure 2 F), (i) both NAP and NRP proteins form their own homo- and heterodimers but they do not interact with each other [ 32 ], (ii) NAP1 does not interact with the CAF-1 (chromatin assembly factor 1) complex subunits (FAS1, FAS2 and MSI1) [ 26 ], and (iii) the ARP4 protein interacts with the RuvBL proteins [ 29 ] providing a link to the known telomerase partner TRB1 [ 15 , 33 ]. RuvBLs and TRB1 were identified among TAP/MS-purified proteins ([ 12 ], Figure 1 A and Figure 2 F).…”

“…Consistently, telomere shortening in fas1/2 , does not seem to occur via interaction with TERT as shown in Jaske et al [ 35 ]. Moreover, the triple mutant fas2nrp1nrp2 retained telomere shortening typical of the fas2 mutant [ 25 ] but in contrast, the quadruple mutants fas1nap1;1nap1;2nap1;3 demonstrated wild-type telomeres [ 26 ]. Altogether these observations could suggest a link to different AtTERT interactions with NAP and NRP proteins that might be coupled with different biological pathways.…”

“…Constructs encoding DOMINO, La1, FIB2, RH2, RH38, LSm4, and HIRP2 were introduced into pSAT1-nEYFP-C1 and/or pSAT1-cEYFP-C1-B Gateway destination vectors [ 76 ]. Protoplasts were isolated from 7-day-old seedlings of Arabidopsis thaliana (Col-0) and transfected as described [ 26 ]. To label cell nuclei and to control transformation efficiency, we co-transfected a plasmid expressing mRFP fused to the nuclear localization signal of the VirD2 protein from Agrobacterium tumefaciens (mRFP-VirD2(NLS); [ 77 ]).…”