Targeted DNA insertion in plants Dong, Oliver Xiaoou; Ronald, Pamela C.
Proceedings of the National Academy of Sciences - PNAS,
06/2021, Letnik:
118, Številka:
22
Journal Article
Recenzirano
Odprti dostop
Conventional methods of DNA sequence insertion into plants, using
Agrobacterium
-mediated transformation or microprojectile bombardment, result in the integration of the DNA at random sites in the ...genome. These plants may exhibit altered agronomic traits as a consequence of disruption or silencing of genes that serve a critical function. Also, genes of interest inserted at random sites are often not expressed at the desired level. For these reasons, targeted DNA insertion at suitable genomic sites in plants is a desirable alternative. In this paper we review approaches of targeted DNA insertion in plant genomes, discuss current technical challenges, and describe promising applications of targeted DNA insertion for crop genetic improvement.
An open source plant kinase chemogenomics set Ercoli, Maria Florencia; Ramos, Priscila Zonzini; Jain, Rashmi ...
Plant direct,
November 2022, Letnik:
6, Številka:
11
Journal Article
Recenzirano
Odprti dostop
One hundred twenty‐nine protein kinases, selected to represent the diversity of the rice (Oryza sativa) kinome, were cloned and tested for expression in Escherichia coli. Forty of these rice kinases ...were purified and screened using differential scanning fluorimetry (DSF) against 627 diverse kinase inhibitors, with a range of structures and activities targeting diverse human kinases. Thirty‐seven active compounds were then tested for their ability to modify primary root development in Arabidopsis. Of these, 14 compounds caused a significant reduction of primary root length compared with control plants. Two of these inhibitory compounds bind to the predicted orthologue of Arabidopsis PSKR1, one of two receptors for PSK, a small sulfated peptide that positively controls root development. The reduced root length phenotype could not be rescued by the exogenous addition of the PSK peptide, suggesting that chemical treatment may inhibit both PSKR1 and its closely related receptor PSKR2. Six of the compounds acting as root growth inhibitors in Arabidopsis conferred the same effect in rice. Compound RAF265 (CHIR‐265), previously shown to bind the human kinase BRAF (B‐Raf proto‐oncogene, serine/threonine kinase), also binds to nine highly conserved rice kinases tested. The binding of human and rice kinases to the same compound suggests that human kinase inhibitor sets will be useful for dissecting the function of plant kinases.
miRNAs contribute to plant resistance against pathogens. Previously, we found that the function of miR398b in immunity in rice differs from that in Arabidopsis. However, the underlying mechanisms are ...unclear. In this study, we characterized the mutants of miR398b target genes and demonstrated that multiple superoxide dismutase genes contribute to miR398b-regulated rice immunity against the blast fungus Magnaporthe oryzae. Out of the four target genes of miR398b, mutations in Cu/Zn-Superoxidase Dismutase1 (CSD1), CSD2 and Os11g09780 (Superoxide DismutaseX, SODX) led to enhanced resistance to M. oryzae and increased hydrogen peroxide (H
O
) accumulation. By contrast, mutations in Copper Chaperone for Superoxide Dismutase (CCSD) resulted in enhanced susceptibility. Biochemical studies revealed that csd1, csd2 and sodx displayed altered expression of CSDs and other superoxide dismutase (SOD) family members, leading to increased total SOD enzyme activity that positively contributed to higher H
O
production. By contrast, the ccsd mutant showed CSD protein deletion, resulting in decreased CSD and total SOD enzyme activity. Our results demonstrate the roles of different SODs in miR398b-regulated resistance to rice blast disease, and uncover an integrative regulatory network in which miR398b boosts total SOD activity to upregulate H
O
concentration and thereby improve disease resistance.
Summary
Nucleotide‐binding leucine‐rich repeat proteins (
NLR
s) serve as intracellular immune receptors in animals and plants. Sensor
NLR
s perceive pathogen‐derived effector molecules and trigger ...robust host defense. Recent studies revealed the role of three coiled‐coil‐type
NLR
s (
CNL
s) of the
ADR
1 family –
ADR
1,
ADR
1‐L1 and
ADR
1‐L2 – as redundant helper
NLR
s, whose function is required for defense mediated by multiple sensor
NLR
s.
From a mutant
snc1
‐enhancing (
MUSE
) forward genetic screen in
A
rabidopsis
targeted to identify negative regulators of
snc1
that encodes a
TIR
‐type
NLR
(
TNL
), we isolated two alleles of
muse15
, both carrying mutations in
ADR
1‐L1
. Interestingly, loss of
ADR
1‐L1
also enhances immunity‐related phenotypes in other autoimmune mutants including
cpr1
,
bal
and
lsd1
. This immunity‐enhancing effect is not mediated by increased
SNC
1 protein stability, nor is it fully dependent on the accumulation of the defense hormone salicylic acid (
SA
).
Transcriptional analysis revealed an upregulation of
ADR
1
and
ADR
1‐L2
in the
adr1‐L1
background, which may overcompensate the loss of
ADR
1‐L1
, resulting in enhanced immunity. Interestingly, autoimmunity of
snc1
and
chs2
, which encode typical
TNL
s, is fully suppressed by the
adr1 triple
mutant, suggesting that the
ADR
s are required for
TNL
downstream signaling.
This study extends our knowledge on the interplay among
ADR
s and reveals their complexity in defense regulation.
Summary
Heat shock proteins (
HSP
s) serve as molecular chaperones for diverse client proteins in many biological processes. In plant immunity, cytosolic
HSP
90s participate in the assembly, ...stability control and/or activation of immune receptor complexes. In this paper we report that in addition to the well‐established positive roles that
HSP
90 isoforms play in plant immunity, they are also involved in the negative regulation of immune receptor accumulation. Point mutations in two
HSP
90
genes,
HSP
90.2
and
HSP
90.3
, were identified from a forward genetic screen designed to isolate mutants with enhanced disease resistance. We found that specific mutations in
HSP
90.2
and
HSP
90.3
lead to heightened accumulation of immune receptors, including
SNC
1,
RPS
2 and
RPS
4.
HSP
90s may assist
SGT
1 in the formation of
SCF
E3 ubiquitin ligase complexes that target immune receptors for degradation. Such regulation is critical for maintaining appropriate levels of immune receptor proteins to avoid autoimmunity.
Nucleocytoplasmic trafficking is emerging as an important aspect of plant immunity. The three related pathways affecting plant immunity include Nuclear Localization Signal (NLS)-mediated nuclear ...protein import, Nuclear Export Signal (NES)-dependent nuclear protein export, and mRNA export relying on MOS3, a nucleoporin belonging to the Nup107-160 complex. Here we report the characterization, identification, and detailed analysis of Arabidopsis modifier of snc1, 11 (mos11). Mutations in MOS11 can partially suppress the dwarfism and enhanced disease resistance phenotypes of snc1, which carries a gain-of-function mutation in a TIR-NB-LRR type Resistance gene. MOS11 encodes a conserved eukaryotic protein with homology to the human RNA binding protein CIP29. Further functional analysis shows that MOS11 localizes to the nucleus and that the mos11 mutants accumulate more poly(A) mRNAs in the nucleus, likely resulting from reduced mRNA export activity. Epistasis analysis between mos3-1 and mos11-1 revealed that MOS11 probably functions in the same mRNA export pathway as MOS3, in a partially overlapping fashion, before the mRNA molecules pass through the nuclear pores. Taken together, MOS11 is identified as a new protein contributing to the transfer of mature mRNA from the nucleus to the cytosol.
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