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Summary Circadian clock, an endogenous time‐setting mechanism, allows plants to adapt to unstable photoperiod conditions and induces flowering with proper timing. In Arabidopsis, the central clock oscillator was formed by a series of interlocked transcriptional feedback loops, but little is known in rice so far. By MutMap technique, we identified the candidate gene OsLHY from a later flowering mutant lem1 and further confirmed it through genetic complementation, RNA interference knockdown, and CRISPR/Cas9‐knockout. Global transcriptome profiling and expression analyses revealed that OsLHY might be a vital circadian rhythm component. Interestingly, oslhy flowered later under ≥12 h day length but headed earlier under ≤11 h day length. qRT‐PCR results exhibited that OsLHY might function through OsGI‐Hd1 pathway. Subsequent one‐hybrid assays in yeast, DNA affinity purification qPCR, and electrophoretic mobility shift assays confirmed OsLHY could directly bind to the CBS element in OsGI promoter. Moreover, the critical day length (CDL) for function reversal of OsLHY in oslhy (11–12 h) was prolonged in the double mutant oslhy osgi (about 13.5 h), indicating that the CDL set by OsLHY was OsGI dependent. Additionally, the dual function of OsLHY entirely relied on Hd1, as the double mutant oslhy hd1 showed the same heading date with hd1 under about 11.5, 13.5, and 14 h day lengths. Together, OsLHY could fine‐tune the CDL by directly regulating OsGI, and Hd1 acts as the final effector of CDL downstream of OsLHY. Our study illustrates a new regulatory mechanism between the circadian clock and photoperiodic flowering.

The iron-sulfur subunit (SDH2) of succinate dehydrogenase plays a key role in electron transport in plant mitochondria. However, it is yet unknown whether SDH2 genes are involved in leaf senescence and yield formation. In this study, we isolated a late premature senescence mutant, lps1, in rice (Oryza sativa). The mutant leaves exhibited brown spots at late tillering stage and wilted at the late grain-filling stage and mature stage. In its premature senescence leaves, photosynthetic pigment contents and net photosynthetic rate were reduced, chloroplasts and mitochondria were degraded. Meanwhile, lps1 displayed small panicles, low seed-setting rate and dramatically reduced grain yield. Gene cloning and complementation analysis suggested that the causal gene for the mutant phenotype was OsSDH2-1 (LOC_Os08g02640), in which single nucleotide mutation resulted in an amino acid substitution in the encoded protein. OsSDH2-1 gene was expressed in all organs tested, with higher expression in leaves, root tips, ovary and anthers. OsSDH2-1 protein was targeted to mitochondria. Furthermore, reactive oxygen species (ROS), mainly H2O2, was excessively accumulated in leaves and young panicles of lps1, which could cause premature leaf senescence and affect panicle development and pollen function. Taken together, OsSDH2-1 plays a crucial role in leaf senescence and yield formation in rice.

Mg-protoporphyrin IX monomethyl ester cyclase (MPEC) plays an essential role in chlorophyll biosynthesis. Further study on the key enzyme will provide us more comprehensive understanding about its function. In the present study, we identified a yellow and spotted leaf 1 (ysl1) mutant in rice exhibiting a yellow-green leaf phenotype throughout the whole developmental stage and a spotted leaf phenotype at seedling and tillering stages. Genetic analysis indicated that a single recessive gene was responsible for the phenotype of ysl1 mutant. Map-based cloning showed that the mutation site was located at the region of 105.4-kb between RM572 and L3 on the short arm of chromosome 1. Sequence analysis revealed that a point mutation (C-to-T) happened in the coding region of candidate gene YSL1 (LOC_Os01g17170) which caused an amino acid change in MPEC. YSL1 was expressed mainly in photosynthetic tissues and organs, and its encoded protein was transported into the chloroplast with a signal peptide. The phenotype of ysl1 returned to normal phenotype of wild type after introducing the wild-type YSL1 gene. Based on the evidence from different levels, we confirmed that the phenotype of ysl1 was caused by a single nucleotide substitution of YSL1 which is allelic to YGL8, OsCRD1, and YL-1, and demonstrated the spotted leaf at seedling and tillering stages as a novel phenotype of the MPEC mutants in rice.

Background Calvin cycle plays a crucial role in carbon fixation which provides the precursors of organic macromolecules for plant growth and development. Currently, no gene involved in Calvin cycle has been identified in monocotyledonous plants through mutant or/and map-based cloning approach. Results Here, we isolated a low-tillering mutant, c6635, in rice (Oryza sativa). The mutant displayed light green leaves and intensely declined pigment contents and photosynthetic capacity at early growth stage. Moreover, its individual plant showed a much smaller size, and most individuals produced only two tillers. At mature stage, its productive panicles, grain number and seed setting rate were significantly decreased, which lead to a sharp reduction of the grain yield. We confirmed that a single nucleotide mutation in LOC_Os04g16680 gene encoding sedoheptulose 1,7-bisphosphatase (SBPase) involved in Calvin cycle was responsible for the mutant phenotype of c6635 through map-based cloning, MutMap analysis and complementation experiments. Sequence analysis suggested that the point mutation caused an amino acid change from Gly-364 to Asp at the C-terminal of SBPase. In addition, OsSBPase gene was mainly expressed in leaf, and the encoded protein was located in chloroplast. The mutation of OsSBPase could significantly affect expression levels of some key genes involved in Calvin cycle. Conclusions We successfully identified a SBPase gene in monocotyledonous plants. Meanwhile, we demonstrated that a single nucleotide substitution at the 3′-end of this gene severely affects plant growth and grain yield, implying that the Gly-364 at the C-terminal of SBPase could play an important role in SBPase function in rice.

Summary: Impairment of microglial clearance activity contributes to beta-amyloid (Aβ) pathology in Alzheimer’s disease (AD). While the transcriptome profile of microglia directs microglial functions, how the microglial transcriptome can be regulated to alleviate AD pathology is largely unknown. Here, we show that injection of interleukin (IL)-33 in an AD transgenic mouse model ameliorates Aβ pathology by reprogramming microglial epigenetic and transcriptomic profiles to induce a microglial subpopulation with enhanced phagocytic activity. These IL-33-responsive microglia (IL-33RMs) express a distinct transcriptome signature that is highlighted by increased major histocompatibility complex class II genes and restored homeostatic signature genes. IL-33-induced remodeling of chromatin accessibility and PU.1 transcription factor binding at the signature genes of IL-33RM control their transcriptome reprogramming. Specifically, disrupting PU.1-DNA interaction abolishes the microglial state transition and Aβ clearance that is induced by IL-33. Thus, we define a PU.1-dependent transcriptional pathway that drives the IL-33-induced functional state transition of microglia, resulting in enhanced Aβ clearance. : Lau et al. show that interleukin-33 (IL-33) enhances microglial amyloid-beta clearance by inducing a subpopulation of MHC-II+ phagocytic microglia, which is, in turn, controlled by PU.1-dependent transcriptome reprogramming. Thus, the authors reveal an IL-33-PU.1 axis involved in transcriptional regulation that promotes beneficial microglial functions in Alzheimer’s disease. Keywords: chemotaxis, phagocytosis, interleukins, beta-amyloid, disease-associated microglia, MHC-II, epigenetics, PU.1, single-cell RNA-sequencing, chromatin accessibility

Abnormal deposition of brain amyloid is a major hallmark of Alzheimer's disease (AD). The toxic extracellular amyloid plaques originating from the aberrant aggregation of beta-amyloid (Aβ) protein are considered to be the major cause of clinical deficits such as memory loss and cognitive impairment. Two-photon excited fluorescence (TPEF) microscopy provides high spatial resolution, minimal invasiveness, and long-term monitoring capability. TPEF imaging of amyloid plaques in AD transgenic mice models has greatly facilitated studies of the AD pathological mechanism. However, the imaging of deep cortical layers is still hampered by the conventional amyloid probes with short excitation/emission wavelength. In this work, we report that a near-infrared (NIR) probe, named CRANAD-3, is far superior for deep in vivo TPEF imaging of brain amyloid in comparison with the commonly used short-wavelength probe. Our findings show that the major interference for TPEF signal of the NIR probe is from the autofluorescence of lipofuscin, the "aging-pigment" in the brain. To eliminate the interference, we characterized the lipofuscin fluorescence in the aged brains of AD mice and found that it has unique broad emission and short lifetime. The lipofuscin signal can be clearly separated from the fluorescence of CRANAD-3 and fluorescent protein via a ratio-based unmixing method. Our results demonstrate the great advantages of NIR probes for in vivo deep-tissue imaging of amyloid plaques in AD.

Amyloid depositions in the brain represent the characteristic hallmarks of Alzheimer’s disease (AD) pathology. The abnormal accumulation of extracellular amyloid-beta (Aβ) and resulting toxic amyloid plaques are considered to be responsible for the clinical deficits including cognitive decline and memory loss. In vivo two-photon fluorescence imaging of amyloid plaques in live AD mouse model through a chronic imaging window (thinned skull or craniotomy) provides a mean to greatly facilitate the study of the pathological mechanism of AD owing to its high spatial resolution and long-term continuous monitoring. However, the imaging depth for amyloid plaques is largely limited to upper cortical layers due to the short-wavelength fluorescence emission of commonly used amyloid probes. In this work, we reported that CRANAD-3, a near-infrared (NIR) probe for amyloid species with excitation wavelength at 900 nm and emission wavelength around 650 nm, has great advantages over conventionally used probes and is well suited for twophoton deep imaging of amyloid plaques in AD mouse brain. Compared with a commonly used MeO-X04 probe, the imaging depth of CRANAD-3 is largely extended for open skull cranial window. Furthermore, by using two-photon excited fluorescence spectroscopic imaging, we characterized the intrinsic fluorescence of the “aging pigment” lipofuscin in vivo, which has distinct spectra from CRANAD-3 labeled plaques. This study reveals the unique potential of NIR probes for in vivo, high-resolution and deep imaging of brain amyloid in Alzheimer’s disease.

Femtosecond laser microsurgery has become an advanced method for clinical procedures and biological research. The tissue treated by femtosecond laser can become highly fluorescent, indicating the formation of new fluorescent compounds that can naturally label the treated tissue site. We systematically characterized the fluorescence signals produced by femtosecond laser ablation in biological tissues in vivo. Our findings showed that they possess unique fluorescence properties and can be clearly differentiated from endogenous signals and major fluorescent proteins. We further demonstrated that the new fluorescent compounds can be used as in vivo labelling agent for biological imaging and guided laser microsurgery.