ABLs and TMKs are co-receptors for extracellular auxin.

Extracellular perception of auxin, an essential phytohormone in plants, has been debated for decades. Auxin-binding protein 1 (ABP1) physically interacts with quintessential transmembrane kinases (TMKs) and was proposed to act as an extracellular auxin receptor, but its role was disputed because abp1 knockout mutants lack obvious morphological phenotypes. Here, we identified two new auxin-binding proteins, ABL1 and ABL2, that are localized to the apoplast and directly interact with the extracellular domain of TMKs in an auxin-dependent manner. Furthermore, functionally redundant ABL1 and ABL2 genetically interact with TMKs and exhibit functions that overlap with those of ABP1 as well as being independent of ABP1. Importantly, the extracellular domain of TMK1 itself binds auxin and synergizes with either ABP1 or ABL1 in auxin binding. Thus, our findings discovered auxin receptors ABL1 and ABL2 having functions overlapping with but distinct from ABP1 and acting together with TMKs as co-receptors for extracellular auxin.

A natural gene drive system confers reproductive isolation in rice.

Hybrid sterility restricts the utilization of superior heterosis of indica-japonica inter-subspecific hybrids. In this study, we report the identification of RHS12, a major locus controlling male gamete sterility in indica-japonica hybrid rice. We show that RHS12 consists of two genes (iORF3/DUYAO and iORF4/JIEYAO) that confer preferential transmission of the RHS12-i type male gamete into the progeny, thereby forming a natural gene drive. DUYAO encodes a mitochondrion-targeted protein that interacts with OsCOX11 to trigger cytotoxicity and cell death, whereas JIEYAO encodes a protein that reroutes DUYAO to the autophagosome for degradation via direct physical interaction, thereby detoxifying DUYAO. Evolutionary trajectory analysis reveals that this system likely formed de novo in the AA genome Oryza clade and contributed to reproductive isolation (RI) between different lineages of rice. Our combined results provide mechanistic insights into the genetic basis of RI as well as insights for strategic designs of hybrid rice breeding.

MicroRNAs inhibit the translation of target mRNAs on the endoplasmic reticulum in Arabidopsis.

Translation inhibition is a major but poorly understood mode of action of microRNAs (miRNAs) in plants and animals. In particular, the subcellular location where this process takes place is unknown. Here, we show that the translation inhibition, but not the mRNA cleavage activity, of Arabidopsis miRNAs requires ALTERED MERISTEM PROGRAM1 (AMP1). AMP1 encodes an integral membrane protein associated with endoplasmic reticulum (ER) and ARGONAUTE1, the miRNA effector and a peripheral ER membrane protein. Large differences in polysome association of miRNA target RNAs are found between wild-type and the amp1 mutant for membrane-bound, but not total, polysomes. This, together with AMP1-independent recruitment of miRNA target transcripts to membrane fractions, shows that miRNAs inhibit the translation of target RNAs on the ER. This study demonstrates that translation inhibition is an important activity of plant miRNAs, reveals the subcellular location of this activity, and uncovers a previously unknown function of the ER.

A century journey of organelles research in the plant endomembrane system.

We are entering an exciting century in the study of the plant organelles in the endomembrane system. Over the past century, especially within the past 50 years, tremendous advancements have been made in the complex plant cell to generate a much clearer and informative picture of plant organelles, including the molecular/morphological features, dynamic/spatial behavior, and physiological functions. Importantly, all these discoveries and achievements in the identification and characterization of organelles in the endomembrane system would not have been possible without: (1) the innovations and timely applications of various state-of-art cell biology tools and technologies for organelle biology research; (2) the continuous efforts in developing and characterizing new organelle markers by the plant biology community; and (3) the landmark studies on the identification and characterization of the elusive organelles. While molecular aspects and results for individual organelles have been extensively reviewed, the development of the techniques for organelle research in plant cell biology is less appreciated. As one of the ASPB Centennial Reviews on "organelle biology," here we aim to take a journey across a century of organelle biology research in plants by highlighting the important tools (or landmark technologies) and key scientists that contributed to visualize organelles. We then highlight the landmark studies leading to the identification and characterization of individual organelles in the plant endomembrane systems.

Autophagosome biogenesis and organelle homeostasis in plant cells.

Autophagy is one of the major highly inducible degradation processes in response to plant developmental and environmental signals. In response to different stimuli, cellular materials, including proteins and organelles, can be sequestered into a double membrane autophagosome structure either selectively or non-selectively. The formation of an autophagosome as well as its delivery into the vacuole involves complex and dynamic membrane processes. The identification and characterization of the conserved autophagy-related (ATG) proteins and their related regulators have greatly advanced our understanding of the molecular mechanism underlying autophagosome biogenesis and function in plant cells. Autophagosome biogenesis is tightly regulated by the coordination of multiple ATG and non-ATG proteins, and selective cargo recruitment. This review updates our current knowledge of autophagosome biogenesis, with special emphasis on the core molecular machinery that drives autophagosome formation, and autophagosome-organelle interactions under abiotic stress conditions.

The plant-unique protein DRIF1 coordinates with sorting nexin 1 to regulate membrane protein homeostasis.

Membrane protein homeostasis is fine-tuned by the cellular pathways for vacuolar degradation and recycling, which ultimately facilitate plant growth and cell-environment interactions. The endosomal sorting complex required for transport (ESCRT) machinery plays important roles in regulating intraluminal vesicle (ILV) formation and membrane protein sorting to vacuoles. We previously showed that the plant-specific ESCRT component FYVE DOMAIN PROTEIN REQUIRED FOR ENDOSOMAL SORTING1 (FREE1) performs multiple functions in plants, although the underlying mechanisms remain elusive. In this study, we performed a suppressor screen of the FREE1-RNAi mutant and identified and characterized 2 suppressor of free1 (sof) mutants in Arabidopsis (Arabidopsis thaliana). These mutants, sof10 and sof641, result in a premature stop codon or a missense mutation in AT5G10370, respectively. This gene was named DEAH and RING domain-containing protein as FREE1 suppressor 1 (DRIF1). DRIF1 has a homologous gene, DRIF2, in the Arabidopsis genome with 95% identity to DRIF1. The embryos of drif1 drif2 mutants arrested at the globular stage and formed enlarged multivesicular bodies (MVBs) with an increased number of ILVs. DRIF1 is a membrane-associated protein that coordinates with retromer component sorting nexin 1 to regulate PIN-FORMED2 recycling to the plasma membrane. Altogether, our data demonstrate that DRIF1 is a unique retromer interactor that orchestrates FREE1-mediated ILV formation of MVBs and vacuolar sorting of membrane proteins for degradation in plants.

Ufmylation reconciles salt stress-induced unfolded protein responses via ER-phagy in Arabidopsis.

In plants, the endomembrane system is tightly regulated in response to environmental stresses for maintaining cellular homeostasis. Autophagosomes, the double membrane organelles forming upon nutrient deprivation or stress induction, degrade bulky cytosolic materials for nutrient turnover. Though abiotic stresses have been reported to induce plant autophagy, few receptors or regulators for selective autophagy have been characterized for specific stresses. Here, we have applied immunoprecipitation followed by tandem mass spectrometry using the autophagosome marker protein ATG8 as bait and have identified the E3 ligase of the ufmylation system Ufl1 as a bona fide ATG8 interactor under salt stress. Notably, core components in the ufmylation cascade, Ufl1 and Ufm1, interact with the autophagy kinase complexes proteins ATG1 and ATG6. Cellular and genetic analysis showed that Ufl1 is important for endoplasmic reticulum (ER)-phagy under persisting salt stress. Loss-of-function mutants of Ufl1 display a salt stress hypersensitive phenotype and abnormal ER morphology. Prolonged ER stress responses are detected in ufl1 mutants that phenocopy the autophagy dysfunction atg5 mutants. Consistently, expression of ufmylation cascade components is up-regulated by salt stress. Taken together, our study demonstrates the role of ufmylation in regulating ER homeostasis under salt stress through ER-phagy.

A plant-unique protein BLISTER coordinates with core retromer to modulate endosomal sorting of plasma membrane and vacuolar proteins.

The retromer is a heteromeric protein complex that localizes to endosomal membranes and drives the formation of endosomal tubules that recycle membrane protein cargoes. In plants, the retromer plays essential and canonical functions in regulating the transport of vacuolar storage proteins and the recycle of endocytosed plasma membrane proteins (PM); however, the mechanisms underlying the regulation of assembly, protein stability, and membrane recruitment of the plant retromer complex remain to be elucidated. In this study, we identify a plant-unique endosomal regulator termed BLISTER (BLI), which colocalizes and associates with the retromer complex by interacting with the retromer core subunits VPS35 and VPS29. Depletion of BLI perturbs the assembly and membrane recruitment of the retromer core VPS26-VPS35-VPS29 trimer. Consequently, depletion of BLI disrupts retromer-regulated endosomal trafficking function, including transport of soluble vacuolar proteins and recycling of endocytosed PIN-FORMED (PIN) proteins from the endosomes back to the PM. Moreover, genetic analysis in Arabidopsis thaliana mutants reveals BLI and core retromer interact genetically in the regulation of endosomal trafficking. Taken together, we identified BLI as a plant-specific endosomal regulator, which functions in retromer pathway to modulate the recycling of endocytosed PM proteins and the trafficking of soluble vacuolar cargoes.

14-3-3 proteins contribute to autophagy by modulating SINAT-mediated degradation of ATG13.

In multicellular eukaryotes, autophagy is a conserved process that delivers cellular components to the vacuole or lysosome for recycling during development and stress responses. Induction of autophagy activates AUTOPHAGY-RELATED PROTEIN 1 (ATG1) and ATG13 to form a protein kinase complex that initiates autophagosome formation. However, the detailed molecular mechanism underlying the regulation of this protein complex in plants remains unclear. Here, we determined that in Arabidopsis thaliana, the regulatory proteins 14-3-3λ and 14-3-3κ redundantly modulate autophagy dynamics by facilitating SEVEN IN ABSENTIA OF ARABIDOPSIS THALIANA (SINAT)-mediated proteolysis of ATG13a and ATG13b. 14-3-3λ and 14-3-3κ directly interacted with SINATs and ATG13a/b in vitro and in vivo. Compared to wild-type (WT), the 14-3-3λ 14-3-3κ double mutant showed increased tolerance to nutrient starvation, delayed leaf senescence, and enhanced starvation-induced autophagic vesicles. Moreover, 14-3-3s were required for SINAT1-mediated ubiquitination and degradation of ATG13a. Consistent with their roles in ATG degradation, the 14-3-3λ 14-3-3κ double mutant accumulated higher levels of ATG1a/b/c and ATG13a/b than the WT upon nutrient deprivation. Furthermore, the specific association of 14-3-3s with phosphorylated ATG13a was crucial for ATG13a stability and formation of the ATG1-ATG13 complex. Thus, our findings demonstrate that 14-3-3λ and 14-3-3κ function as molecular adaptors to regulate autophagy by modulating the homeostasis of phosphorylated ATG13.

The plant ESCRT component FREE1 regulates peroxisome-mediated turnover of lipid droplets in germinating Arabidopsis seedlings.

Lipid droplets (LDs) stored during seed development are mobilized and provide essential energy and lipids to support seedling growth upon germination. Triacylglycerols (TAGs) are the main neutral lipids stored in LDs. The lipase SUGAR DEPENDENT 1 (SDP1), which hydrolyzes TAGs in Arabidopsis thaliana, is localized on peroxisomes and traffics to the LD surface through peroxisomal extension, but the underlying mechanism remains elusive. Here, we report a previously unknown function of a plant-unique endosomal sorting complex required for transport (ESCRT) component FYVE DOMAIN PROTEIN REQUIRED FOR ENDOSOMAL SORTING 1 (FREE1) in regulating peroxisome/SDP1-mediated LD turnover in Arabidopsis. We showed that LD degradation was impaired in germinating free1 mutant; moreover, the tubulation of SDP1- or PEROXIN 11e (PEX11e)-marked peroxisomes and the migration of SDP1-positive peroxisomes to the LD surface were altered in the free1 mutant. Electron tomography analysis showed that peroxisomes failed to form tubules to engulf LDs in free1, unlike in the wild-type. FREE1 interacted directly with both PEX11e and SDP1, suggesting that these interactions may regulate peroxisomal extension and trafficking of the lipase SDP1 to LDs. Taken together, our results demonstrate a pivotal role for FREE1 in LD degradation in germinating seedlings via regulating peroxisomal tubulation and SDP1 targeting.

Arabidopsis ORP2A mediates ER-autophagosomal membrane contact sites and regulates PI3P in plant autophagy.

Autophagy is an intracellular degradation system for cytoplasmic constituents which is mediated by the formation of a double-membrane organelle termed the autophagosome and its subsequent fusion with the lysosome/vacuole. The formation of the autophagosome requires membrane from the endoplasmic reticulum (ER) and is tightly regulated by a series of autophagy-related (ATG) proteins and lipids. However, how the ER contacts autophagosomes and regulates autophagy remain elusive in plants. In this study, we identified and demonstrated the roles of Arabidopsis oxysterol-binding protein-related protein 2A (ORP2A) in mediating ER-autophagosomal membrane contacts and autophagosome biogenesis. We showed that ORP2A localizes to both ER-plasma membrane contact sites (EPCSs) and autophagosomes, and that ORP2A interacts with both the ER-localized VAMP-associated protein (VAP) 27-1 and ATG8e on the autophagosomes to mediate the membrane contact sites (MCSs). In ORP2A artificial microRNA knockdown (KD) plants, seedlings display retarded growth and impaired autophagy levels. Both ATG1a and ATG8e accumulated and associated with the ER membrane in ORP2A KD lines. Moreover, ORP2A binds multiple phospholipids and shows colocalization with phosphatidylinositol 3-phosphate (PI3P) in vivo. Taken together, ORP2A mediates ER-autophagosomal MCSs and regulates autophagy through PI3P redistribution.

Plant ESCRT protein ALIX coordinates with retromer complex in regulating receptor-mediated sorting of soluble vacuolar proteins.

Vacuolar proteins play essential roles in plant physiology and development, but the factors and the machinery regulating their vesicle trafficking through the endomembrane compartments remain largely unknown. We and others have recently identified an evolutionarily conserved plant endosomal sorting complex required for transport (ESCRT)-associated protein apoptosis-linked gene-2 interacting protein X (ALIX), which plays canonical functions in the biogenesis of the multivesicular body/prevacuolar compartment (MVB/PVC) and in the sorting of ubiquitinated membrane proteins. In this study, we elucidate the roles and underlying mechanism of ALIX in regulating vacuolar transport of soluble proteins, beyond its conventional ESCRT function in eukaryotic cells. We show that ALIX colocalizes and physically interacts with the retromer core subunits Vps26 and Vps29 in planta. Moreover, double-mutant analysis reveals the genetic interaction of ALIX with Vps26 and Vps29 for regulating trafficking of soluble vacuolar proteins. Interestingly, depletion of ALIX perturbs membrane recruitment of Vps26 and Vps29 and alters the endosomal localization of vacuolar sorting receptors (VSRs). Taken together, ALIX functions as a unique retromer core subcomplex regulator by orchestrating receptor-mediated vacuolar sorting of soluble proteins.

Structural insights into how vacuolar sorting receptors recognize the sorting determinants of seed storage proteins.

In Arabidopsis, vacuolar sorting receptor isoform 1 (VSR1) sorts 12S globulins to the protein storage vacuoles during seed development. Vacuolar sorting is mediated by specific protein-protein interactions between VSR1 and the vacuolar sorting determinant located at the C terminus (ctVSD) on the cargo proteins. Here, we determined the crystal structure of the protease-associated domain of VSR1 (VSR1-PA) in complex with the C-terminal pentapeptide (468RVAAA472) of cruciferin 1, an isoform of 12S globulins. The 468RVA470 motif forms a parallel ß-sheet with the switch III residues (127TMD129) of VSR1-PA, and the 471AA472 motif docks to a cradle formed by the cargo-binding loop (95RGDCYF100), making a hydrophobic interaction with Tyr99. The C-terminal carboxyl group of the ctVSD is recognized by forming salt bridges with Arg95. The C-terminal sequences of cruciferin 1 and vicilin-like storage protein 22 were sufficient to redirect the secretory red fluorescent protein (spRFP) to the vacuoles in Arabidopsis protoplasts. Adding a proline residue to the C terminus of the ctVSD and R95M substitution of VSR1 disrupted receptor-cargo interactions in vitro and led to increased secretion of spRFP in Arabidopsis protoplasts. How VSR1-PA recognizes ctVSDs of other storage proteins was modeled. The last three residues of ctVSD prefer hydrophobic residues because they form a hydrophobic cluster with Tyr99 of VSR1-PA. Due to charge-charge interactions, conserved acidic residues, Asp129 and Glu132, around the cargo-binding site should prefer basic residues over acidic ones in the ctVSD. The structural insights gained may be useful in targeting recombinant proteins to the protein storage vacuoles in seeds.

Embryo sac development relies on symplastic signals from ovular integuments in Arabidopsis.

Ovules are female reproductive organs of angiosperms, consisting of sporophytic integuments surrounding female gametophytes, that is, embryo sacs. Synchronization between integument growth and embryo sac development requires intracellular communication. However, signaling routes through which cells of the two generations communicate are unclear. We report that symplastic signals through plasmodesmata (PDs) of integuments are critical for the development of female gametophytes. Genetic interferences of PD biogenesis either by functional loss of CHOLINE TRANSPORTER-LIKE1 (CTL1) or by integument-specific expression of a mutated CALLOSE SYNTHASE 3 (cals3m) compromised PD formation in integuments and reduced fertility. Close examination of pINO:cals3m or ctl1 ovules indicated that female gametophytic development was either arrested at various stages after the formation of functional megaspores. In both cases, defective ovules could not attract pollen tubes, leading to the failure of fertilization. Results presented here demonstrate a key role of the symplastic route in sporophytic control of female gametophytic development.

Arabidopsis AUTOPHAGY-RELATED2 is essential for ATG18a and ATG9 trafficking during autophagosome closure.

As a fundamental metabolic pathway, autophagy plays important roles in plant growth and development, particularly under stress conditions. A set of autophagy-related (ATG) proteins is recruited for the formation of a double-membrane autophagosome. Among them, the essential roles of ATG2, ATG18, and ATG9 have been well established in plant autophagy via genetic analysis; however, the underlying molecular mechanism for ATG2 in plant autophagosome formation remains poorly understood. In this study, we focused on the specific role of ATG2 in the trafficking of ATG18a and ATG9 during autophagy in Arabidopsis (Arabidopsis thaliana). Under normal conditions, YFP-ATG18a proteins are partially localized on late endosomes and translocated to ATG8e-labeled autophagosomes upon autophagic induction. Real-time imaging analysis revealed sequential recruitment of ATG18a on the phagophore membrane, showing that ATG18a specifically decorated the closing edges and finally disassociated from the completed autophagosome. However, in the absence of ATG2, most of the YFP-ATG18a proteins are arrested on autophagosomal membranes. Ultrastructural and 3D tomography analysis showed that unclosed autophagosome structures are accumulated in the atg2 mutant, displaying direct connections with the endoplasmic reticulum membrane and vesicular structures. Dynamic analysis of ATG9 vesicles suggested that ATG2 depletion also affects the association between ATG9 vesicles and the autophagosomal membrane. Furthermore, using interaction and recruitment analysis, we mapped the interaction relationship between ATG2 and ATG18a, implying a possible role of ATG18a in recruiting ATG2 and ATG9 to the membrane. Our findings unveil a specific role of ATG2 in coordinating ATG18a and ATG9 trafficking to mediate autophagosome closure in Arabidopsis.

Frequency-domain terahertz optoacoustics for non-contact quantitative detection of gas, liquid, and solid samples.

Terahertz optoacoustics (THz-OA) combines the advantages of abundant molecular characteristic absorptions in a terahertz band and the low attenuation through ultrasonic detection. Frequency-domain THz-OA, benefiting from the compact and the low cost of a continuous-wave THz source, has been used in gas detection and sensing. However, liquid and solid detections are hard to achieve due to the sensitivity limitation of existing technologies. Here we present a high-sensitivity frequency-domain THz-OA system with customized optoacoustic cells to accomplish non-contact quantitative detection of gas, liquid, and solid samples. The relationships between signal amplitudes and sample concentration, volume and temperature are discussed separately, revealing a potential application of this technology.

An in vitro vesicle formation assay reveals cargo clients and factors that mediate vesicular trafficking.

The fidelity of protein transport in the secretory pathway relies on the accurate sorting of proteins to their correct destinations. To deepen our understanding of the underlying molecular mechanisms, it is important to develop a robust approach to systematically reveal cargo proteins that depend on specific sorting machinery to be enriched into transport vesicles. Here, we used an in vitro assay that reconstitutes packaging of human cargo proteins into vesicles to quantify cargo capture. Quantitative mass spectrometry (MS) analyses of the isolated vesicles revealed cytosolic proteins that are associated with vesicle membranes in a GTP-dependent manner. We found that two of them, FAM84B (also known as LRAT domain containing 2 or LRATD2) and PRRC1, contain proline-rich domains and regulate anterograde trafficking. Further analyses revealed that PRRC1 is recruited to endoplasmic reticulum (ER) exit sites, interacts with the inner COPII coat, and its absence increases membrane association of COPII. In addition, we uncovered cargo proteins that depend on GTP hydrolysis to be captured into vesicles. Comparing control cells with cells depleted of the cargo receptors, SURF4 or ERGIC53, we revealed specific clients of each of these two export adaptors. Our results indicate that the vesicle formation assay in combination with quantitative MS analysis is a robust and powerful tool to uncover novel factors that mediate vesicular trafficking and to uncover cargo clients of specific cellular factors.

A unique AtSar1D-AtRabD2a nexus modulates autophagosome biogenesis in Arabidopsis thaliana.

In eukaryotes, secretory proteins traffic from the endoplasmic reticulum (ER) to the Golgi apparatus via coat protein complex II (COPII) vesicles. Intriguingly, during nutrient starvation, the COPII machinery acts constructively as a membrane source for autophagosomes during autophagy to maintain cellular homeostasis by recycling intermediate metabolites. In higher plants, essential roles of autophagy have been implicated in plant development and stress responses. Nonetheless, the membrane sources of autophagosomes, especially the participation of the COPII machinery in the autophagic pathway and autophagosome biogenesis, remains elusive in plants. Here, we provided evidence in support of a novel role of a specific Sar1 homolog AtSar1d in plant autophagy in concert with a unique Rab1/Ypt1 homolog AtRabD2a. First, proteomic analysis of the plant ATG (autophagy-related gene) interactome uncovered the mechanistic connections between ATG machinery and specific COPII components including AtSar1d and Sec23s, while a dominant negative mutant of AtSar1d exhibited distinct inhibition on YFP-ATG8 vacuolar degradation upon autophagic induction. Second, a transfer DNA insertion mutant of AtSar1d displayed starvation-related phenotypes. Third, AtSar1d regulated autophagosome progression through specific recognition of ATG8e by a noncanonical motif. Fourth, we demonstrated that a plant-unique Rab1/Ypt1 homolog AtRabD2a coordinates with AtSar1d to function as the molecular switch in mediating the COPII functions in the autophagy pathway. AtRabD2a appears to be essential for bridging the specific AtSar1d-positive COPII vesicles to the autophagy initiation complex and therefore contributes to autophagosome formation in plants. Taken together, we identified a plant-specific nexus of AtSar1d-AtRabD2a in regulating autophagosome biogenesis.

SOX9-COL9A3-dependent regulation of choroid plexus epithelial polarity governs blood-cerebrospinal fluid barrier integrity.

The choroid plexus (CP) is an extensively vascularized neuroepithelial tissue that projects into the brain ventricles. The restriction of transepithelial transport across the CP establishes the blood-cerebrospinal fluid (CSF) barrier that is fundamental to the homeostatic regulation of the central nervous system microenvironment. However, the molecular mechanisms that control this process remain elusive. Here we show that the genetic ablation of Sox9 in the hindbrain CP results in a hyperpermeable blood-CSF barrier that ultimately upsets the CSF electrolyte balance and alters CSF protein composition. Mechanistically, SOX9 is required for the transcriptional up-regulation of Col9a3 in the CP epithelium. The reduction of Col9a3 expression dramatically recapitulates the blood-CSF barrier defects of Sox9 mutants. Loss of collagen IX severely disrupts the structural integrity of the epithelial basement membrane in the CP, leading to progressive loss of extracellular matrix components. Consequently, this perturbs the polarized microtubule dynamics required for correct orientation of apicobasal polarity and thereby impedes tight junction assembly in the CP epithelium. Our findings reveal a pivotal cascade of SOX9-dependent molecular events that is critical for construction of the blood-CSF barrier.

Fully Dispersed IrOx Atomic Clusters Enable Record Photoelectrochemical Water Oxidation of Hematite in Acidic Media.

Ir-based materials are still the benchmark catalysts for various reactions in acidic environment, but the high loading and low atom utilization limit their large-scale deployment. Herein, we report an effective strategy for implanting fully dispersed iridium-oxide atomic clusters onto hematite for boosting photoelectrochemical water oxidation in acidic solution. The resulting photoanode achieves a record-high photocurrent of 1.35 mA cm-2 at 1.23 V, corresponding to a mass activity of 172.70 A g-1 (3 times higher than electrodeposited control sample) and demonstrating the merits from the high atomic utilization of Ir. The systematically experimental and theoretical results reveal that the performance improvement correlates with the modulated electronic structure including the adjusted Fermi level and d-band center, which significantly enhances charge separation efficiency and promotes the conversion from intermediate *O into *OOH.