水稻籼粳杂种生殖障碍的基因定位分析

籼稻(Oryza sativa L.ssp.indica)与粳稻(O.sativa ssp.japonica)杂交优势明显但存在生殖隔离。生殖障碍主要表现为胚囊败育、花粉败育、开花时花药不开裂和雌雄异熟。应用具有137个标记位点的籼、粳杂交(“窄叶青8号”/“京系17”)F_1花药培养获得的127个双单倍体(DH)群体构建的RFLP图谱,对控制籼、粳杂种小穗败育的基因座位进行了定位研究。结果在第1、3、4、5、6、7、8、12染色体上检测到10个基因座位,其中第3、12染色体上的2个不育基因位点stj-3和stj-12与同一杂交组合F_2分离群体中发现的异常分离热点处于相同的染色体区段。Ssj-6的基因加性效应为负值,有增加籼、粳亲和性的作用;其余的不育基因座位皆有增加籼、粳杂种不育性的作用。     

QTLs for hybrid fertility and their association with female and male sterility in rice

Hybrid sterility is one of the major barriers to the application of wide crosses in plant breeding and is commonly encountered in crosses between indica and japonica rice varieties. Ten mapping populations comprised of two reciprocal F2 and eight BC₁F₁ populations generated from the cross between Ilpumbyeo (japonica) and Dasanbyeo (indica) were used to identify QTLs and to interpret the gametophytic factors involved in hybrid fertility or sterility between two subspecies. Frame maps were constructed using a total of 107 and 144 STS markers covering 12 rice chromosomes in two reciprocal F2 and eight BC₁F₁ populations, respectively. A total of 15 main-effect QTLs and 17 significant digenic-epistatic interactions controlling spikelet fertility (SF) were resolved in the entire genome map of F₂ and BC₁F₁ populations. Among detected QTLs responsible for hybrid fertility, four QTLs, qSF5.1 and qSF5.2 on chromosome 5, qSF6.2 on chromosome 6, and qSF12.2 on chromosome 12, were identified as major QTLs since they were located at corresponding positions in at least three mapping populations. Loci qSF5.1, qSF6.1 and qSF6.2 were responsible for both female and male sterility, whereas qSF3.1, qSF7 and qSF12.2 affected the spikelet fertility only through embryosac factors, and qSF9.1 did through pollen factors. Five new QTLs identified in this study will be helpful for understanding the hybrid sterility and for breeding programs via inter-subspecific hybridization.     

Genetic dissection of embryo sac fertility, pollen fertility, and their contributions to spikelet fertility of intersubspecific hybrids in rice

The partial sterility of hybrids has been a major barrier for utilization of the strong heterosis expressed in hybrids between Oryza sativa ssp. indica and O. sativa ssp. japonica. Wide-compatibility varieties, comprising a special class of germplasm, are able to produce fertile hybrids when crossed to both indica and japonica varieties. However, all the work on wide compatibility and majority of studies on indica/japonica hybrid sterility reported so far were based only on spikelet fertility; thus, it is not known to what extent male and female gamete abortions influence hybrid sterility. In this study, we investigated pollen fertility, embryo sac fertility, and spikelet fertility in an F₁ population of 202 true hybrid plants derived from a three-way cross (02428/Nanjing 11//Balilla). A partial regression analysis showed that the pollen and embryo sac fertility contributed almost equally to spikelet fertility. QTL analysis based on a linkage map of 191 polymorphic marker loci identified two QTLs for pollen fertility, one QTL for embryo sac fertility, and three QTLs for spikelet fertility. The S5 locus, previously identified as a locus for wide compatibility by spikelet fertility analysis, is a major locus for embryo sac fertility, and a QTL on chromosome 5 had a major effect on pollen fertility. These two loci coincided with the two major QTLs for spikelet fertility. The study also detected a QTL on chromosome 8, showing a large effect on spikelet fertility but no effect on either pollen or embryo sac fertility. Very little interaction among the QTLs was detected. The implications of the findings in rice breeding programs are discussed.     

Identification for quantitative trait loci controlling grain shattering in rice

Seed shattering is an important factor causing loss of grain yield before and during rice harvest. In the present study, the quantitative trait loci regarding shattering scale, breaking tensile strength (BTS) and abscission layer (AL), the parameters evaluating seed shattering habit by hand gripping, a digital force gauge and observation on AL, respectively, were identified by using an doubled haploid line (DHL) population from a cross between a loose-shattering type Tongil variety, ‘Samgang’, and a moderately difficult shattering japonica variety, ‘Nagdong’. Eight QTLs consisted in four QTLs for shattering scale, two QTLs for AL, each one QTL for pulling and bending strength were detected on six chromosomes, respectively. Among them, Qss1 with flanking markers RM6696 and RM476 explained 31% of phenotype variation in shattering scale. Furthermore, two new QTLs controlling shattering habit, Qss5-2 and Qal5-1, were located on chromosome 5 at the interval 5028–5037 and 5021-RM289. They explained 10% and 12% of phenotype variations, respectively. A total of eleven digenic epistatic loci were identified for four parameters. The identification of QTLs affecting seed shattering habits is favorable to thoroughly dissect the genetic mechanism of the shattering habit and to apply for marker-assisted selection in rice breeding system of specific regions.     

Increasing the accuracy of genomic prediction in pure‐bred Limousin beef cattle by including cross‐bred Limousin data and accounting for an F94L variant in MSTN

Explicitly fitting effects for major genes or QTL that account for a large percentage of variation in a whole genomic prediction model may increase prediction accuracy. This study compared approaches to account for a major effect of an F94L variant in the MSTN gene within the genomic prediction using bovine whole‐genomic SNP markers. Among the beef cattle breeds, Limousin have been known to have an F94L variant that is not present in Angus. The reference population in this study consisted of 3060 beef cattle including pure‐bred Limousin (PL), cross‐bred Limousin with Angus (LF) and pure‐bred Angus, genotyped using a BovineSNP50 BeadChip and directly for the MSTN‐F94L variant. We compared prediction accuracies in PL animals using the three datasets from only the PL population, admixed PL and LF (AL) or multibreed analysis using all of the PL, LF and Angus (MB) population according to four‐fold cross‐validation after K‐means clustering. The MSTN‐F94L variant was the most strongly associated with five traits (birth weight, calving ease direct, milk, weaning weight and yield grade) among the 13 measured traits in PL and AL populations. Fitting the MSTN‐F94L variant as a random effect, the genomic prediction accuracies for birth weight increased by 2.7% in PL, by 2.2% in AL and by 3.2% in MB. Prediction accuracies for five traits increased in the MB analysis. Fitting MSTN‐F94L as a fixed effect in PL, AL and MB analyses resulted in increased prediction accuracy in PL for eight traits. Prediction accuracies can be improved by including a causal variant in genomic evaluation compared with simply using whole‐genome SNP markers. Fitting the causal variant as a fixed effect along with markers fitted as random effects resulted in greater prediction accuracies for most traits. Causal variants should be genotyped along with SNP markers.     

d‐glufosinate as a male sterility agent for hybrid seed production

A chemical male sterility system based on anther‐localized conversion of the inactive d ‐enantiomer of the herbicide, glufosinate (2‐amino‐4‐(methylphosphinyl)‐butanoate) to the phytotoxic l is described. Highly pure d ‐glufosinate was isolated in >98% enantiomeric excess from the racemate via fermentation with a strain of Escherichia coli expressing the PAT (l ‐glufosinate N‐acetyl transferase) gene and purification of the unreacted d ‐enantiomer from the broth by ion exchange. A modified (F58K, M213S) form of the d ‐amino acid oxidase (DAAO) (EC 1.4.3.3) from Rhodosporidium toruloides was designed, tested in vitro and found to efficiently oxidize d ‐glufosinate to its 2‐oxo derivative [2‐oxo‐4‐(methylphosphinyl)‐butanoic acid]. Tobacco (Nicotiana tabacum) plants were transformed to express this modified oxidase under control of the TAP1 tapetum‐specific promoter. A number of the resultant transgenic lines exhibited complete male sterility that persisted for two or more weeks immediately following foliar treatment with 75 or 200 g/ha of d ‐glufosinate without exhibiting obvious phytotoxic symptoms or any measurable decline in female fertility. Similarly, plants containing the same construct and, additionally, a PAT gene expressed from a plastocyanin promoter exhibited significantly reduced male fertility and no reduction in female fertility following foliar application of racemic glufosinate. Thus, foliar application of d ‐glufosinate either purified or as the commercial herbicide, combined with anther expression of a modified DAAO promises to provide a cost‐effective conditional chemical male sterility system with the characteristics necessary for practical F₁ hybrid seed production.     

RFLP mapping of QTLs for yield and related characters in rice (Oryza sativa L.)

Quantitative triat loci (QTLs) for yield and related traits in rice were mapped based on RFLP maps from two indica/indica F₂ populations, Tesanai 2/CB and Waiyin 2/CB. In Tesanai 2/CB, 14 intervals carrying QTLs for eight traits were detected, including 3 for grain weight per plant (GWT), 2 for number of panicles per plant (NP), 2 for number of grains per panicle (NG), 1 for total number of spikelets per panicle (TNS), 1 for spikelet fertility (SF), 3 for 1000-grain weight (TGWT), 1 for spikelet density (SD), and 1 for number of first branches per main panicle. The 3 QTLs for GWT were located on chromosomes 1, 2, and 4, with 1 in each chromosome. The additive effect of the single locus ranged from 2.0 g to 9.1 g. A major gene (np4) for NP was detected on chromosome 4 within the interval of RG143–RG214, about 4cM for RG143, and this locus explained 26.1% of the observed phenotypic variance for NP. The paternal allele of this locus was responsible for reduced panicles per plant (3 panicles per plant). In another population, Waiyin 2/CB, 12 intervals containing QTLs for six of the above-mentioned traits were detected, including 3 for GWT, 2 for each of NP, TNS, TGWT and SD, 1 for SF. Three QTLs for GWT were located on chromosome 1, 4, and 5, respectively. The additive effect of the single locus for GWT ranged from 6.7 g to 8.8 g, while the dominance effect was 1.7–11.5 g. QTL mapping in two populations with a common male parent is compared and discussed.     

Three representative inter and intra‐subspecific crosses reveal the genetic architecture of reproductive isolation in rice

Systematic characterization of genetic and molecular mechanisms in the formation of hybrid sterility is of fundamental importance in understanding reproductive isolation and speciation. Using ultra‐high‐density genetic maps, 43 single‐locus quantitative trait loci (QTLs) and 223 digenic interactions for embryo‐sac, pollen, and spikelet fertility are depicted from three crosses between representative varieties of japonica and two varietal groups of indica, which provide an extensive archive for investigating the genetic basis of reproductive isolation in rice. Ten newly detected single‐locus QTLs for inter‐ and intra‐subspecific fertility are identified. Three loci for embryo‐sac fertility are detected in both Nip × ZS97 and Nip × MH63 crosses, whereas QTLs for pollen fertility are not in common between the two crosses thus leading to fertility variation. Five loci responsible for fertility and segregation distortion are observed in the ZS97 × MH63 cross. The importance of two‐locus interactions on fertility are quantified in the whole genome, which identify that three types of interaction contribute to fertility reduction in the hybrid. These results construct the genetic architecture with respect to various forms of reproductive barriers in rice, which have significant implications in utilization of inter‐subspecific heterosis along with improvement in the fertility of indica–indica hybrids at single‐ and multi‐locus level.     

Mapping segregation distortion loci and quantitative trait loci for spikelet sterility in rice ( Oryza sativa L.)

Markers with segregation ratio distortion are commonly observed in data sets used for quantitative trait locus (QTL) mapping. In this study, a multipoint method of maximum likelihood (ML) was newly developed to estimate the positions and effects of the segregation distortion loci (SDLs) in two F2 populations of rice (Oryza sativa L.), i.e. Taichung65/Bhadua (TB; japonica-indica cross) and CPSLO17/W207-2 (CW; japonica-japonica). Of the four parents, W207-2 and Bhadua were found to be spikelet semi-sterile and stably inherited through selfing, and spikelet fertility segregated in the two populations. Therefore, recombination frequencies were recalculated after mapping the SDLs by using the multipoint method, and the molecular linkage maps of the two F2 populations were constructed to detect QTLs underlying spikelet fertility. As a result, five SDLs in the TB population were mapped on chromosomes 1, 3, 8 and 9, respectively. Two major QTLs underlying spikelet fertility, namely qSS-6a and qSS-8a, were detected on chromosomes 6 and 8, respectively. In the CW population, a total of 12 SDLs were detected on all 12 chromosomes except 1, 5, 7 and 11. Three QTLs underlying spikelet sterility, namely qSS-2, qSS-6b and qSS-8b on chromosomes 2, 6 and 8, were determined on the whole genome scale. Interestingly, both qSS-6a and qSS-6b, detected in the two F2 populations respectively, were located on a similar position as the S5 gene on chromosome 6; while qSS-8a and qSS-8b were also simultaneously detected on similar positions of the short arm of chromosome 8 in the two populations, which should be a new sterility gene showing the same type of zygotic selection.     

Genetic dissection of pre‐anthesis sub‐phase durations during the reproductive spike development of wheat

Flowering time is an important factor affecting grain yield in wheat. In this study, we divided reproductive spike development into eight sub‐phases. These sub‐phases have the potential to be delicately manipulated to increase grain yield. We measured 36 traits with regard to sub‐phase durations, determined three grain yield‐related traits in eight field environments and mapped 15 696 single nucleotide polymorphism (SNP, based on 90k Infinium chip and 35k Affymetrix chip) markers in 210 wheat genotypes. Phenotypic and genetic associations between grain yield traits and sub‐phase durations showed significant consistency (Mantel test; r = 0.5377, P < 0.001). The shared quantitative trait loci (QTLs) revealed by the genome‐wide association study suggested a close association between grain yield and sub‐phase duration, which may be attributed to effects on spikelet initiation/spikelet number (double ridge to terminal spikelet stage, DR‐TS) and assimilate accumulation (green anther to anthesis stage, GA‐AN). Moreover, we observed that the photoperiod‐sensitivity allele at the Ppd‐D1 locus on chromosome 2D markedly extended all sub‐phase durations, which may contribute to its positive effects on grain yield traits. The dwarfing allele at the Rht‐D1 (chromosome 4D) locus altered the sub‐phase duration and displayed positive effects on grain yield traits. Data for 30 selected genotypes (from among the original 210 genotypes) in the field displayed a close association with that from the greenhouse. Most importantly, this study demonstrated specific connections to grain yield in narrower time windows (i.e. the eight sub‐phases), rather than the entire stem elongation phase as a whole.     

CHROMOSOMAL REARRANGEMENTS AND THE GENETICS OF REPRODUCTIVE BARRIERS IN MIMULUS (MONKEY FLOWERS)

Chromosomal rearrangements may directly cause hybrid sterility and can facilitate speciation by preserving local adaptation in the face of gene flow. We used comparative linkage mapping with shared gene‐based markers to identify potential chromosomal rearrangements between the sister monkeyflowers Mimulus lewisii and Mimulus cardinalis, which are textbook examples of ecological speciation. We then remapped quantitative trait loci (QTLs) for floral traits and flowering time (premating isolation) and hybrid sterility (postzygotic isolation). We identified three major regions of recombination suppression in the M. lewisii × M. cardinalis hybrid map compared to a relatively collinear Mimulus parishii × M. lewisii map, consistent with a reciprocal translocation and two inversions specific to M. cardinalis. These inferences were supported by targeted intraspecific mapping, which also implied a M. lewisii‐specific reciprocal translocation causing chromosomal pseudo‐linkage in both hybrid mapping populations. Floral QTLs mapped in this study, along with previously mapped adaptive QTLs, were clustered in putatively rearranged regions. All QTLs for male sterility, including two underdominant loci, mapped to regions of recombination suppression. We argue that chromosomal rearrangements may have played an important role in generating and consolidating barriers to gene flow as natural selection drove the dramatic ecological and morphological divergence of these species.     

Epidermal jasmonate perception is sufficient for all aspects of jasmonate‐mediated male fertility in Arabidopsis

Jasmonate (JA) signaling is essential for several environmental responses and reproductive development in many plant species. In Arabidopsis thaliana, the most obvious phenotype of JA biosynthetic and perception mutants is profound sporophytic male sterility characterized by failure of stamen filament elongation, severe delay of anther dehiscence and pollen inviability. The site of action of JA in the context of reproductive development has been discussed, but the ideas have not been tested experimentally. To this end we used targeted expression of a COI1‐YFP transgene in the coi1‐1 mutant background. As COI1 is an essential component of the JA co‐receptor complex, the null coi1‐1 mutant is male sterile due to lack of JA perception. We show that expression of COI1‐YFP in the epidermis of the stamen filament and anther in coi1 mutant plants is sufficient to rescue filament elongation, anther dehiscence and pollen viability. In contrast, filament expression alone or expression in the tapetum do not restore dehiscence and pollen viability. These results demonstrate that epidermal JA perception is sufficient for anther function and pollen viability, and suggest the presence of a JA‐dependent non‐autonomous signal produced in the anther epidermis to synchronize both anther dehiscence and pollen maturation.     

Retracted: Screening of candidate genes and fine mapping of drought tolerance quantitative trait loci on chromosome 4 in rice (Oryza sativa L.) under drought stress

Due to severe water resource shortage, genetics of and breeding for DT (drought tolerance) in rice (Oryza sativa L.) have become one of the hot research topics. Identification of grain yield QTLs (quantitative trait loci) directly related to the DT trait of rice can provide useful information for breeding new drought‐resistant and water‐saving rice varieties via marker‐assisted selection. A population of 105 advanced BILs (backcross introgression lines) derived from a cross between Zhenshan97B and IRAT109 in Zhenshan97B background were grown under drought stress in a field experiment and phenotypic traits were investigated. The results showed that in the target interval of RM273‐RM255 on chromosome 4, three main‐effect QTLs related to panicle length, panicle number, and spikelet number per panicle were identified (LOD [logarithm of the odds] > 2.0). The panicle length‐related QTL had two loci located in the neighboring intervals of RM17308‐RM17305 and RM17349‐RM17190, which explained 18.80% and 20.42%, respectively, of the phenotypic variation, while the panicle number‐related QTL was identified in the interval of RM1354‐RM17308, explaining 11.47% of the phenotypic variation. As far as the spikelet number per panicle‐related QTL was concerned, it was found to be located in the interval of RM17308‐RM17305, which explained 28.08% of the phenotypic variation. Using the online Plant‐GE query system, a total of 13 matched ESTs (expressed sequence tags) were found in the target region, and of the 13 ESTs, 12 had corresponding predicted genes. For instance, the two ESTs CB096766 and CA765747 were corresponded to the same predicted gene LOC_Os04g46370, while the other four ESTs, CA754286, CB000011, CX056247, and CX056240, were corresponded to the same predicted gene LOC_Os04g46390.     

Winter hardiness of Miscanthus (II): Genetic mapping for overwintering ability and adaptation traits in three interconnected Miscanthus populations

Miscanthus × giganteus (M×g) is the primary species of Miscanthus for bioenergy feedstock production. The current leading biomass cultivar, M×g ‘1993‐1780’, is insufficiently adapted in temperate regions with cold winters such as USDA hardiness zone 5 (average annual minimum temperature of ?28.9 to ?23.3°C) or lower. Three interconnected Miscanthus F₁ populations that shared a common parent were planted in a replicated field trial at Urbana, IL (hardiness zone 5b; average annual minimum temperature of ?26.1 to ?23.3°C) in spring 2011. The winter of 2013–2014 was especially cold in Urbana, with a minimum soil temperature at 10 cm of ?6.2°C and a minimum air temperature of ?25.3°C, giving us an opportunity to evaluate hardiness on established year‐3 plants. The parent in common to all three populations, M. sinensis ssp. condensatus ‘Cosmopolitan’, is native to maritime southern Japan, and in Urbana, it is winter‐damaged most years. In contrast, the three other parents, M. sacchariflorus ‘Robustus’ (MapA), M. sinensis ‘Silberturm’ (MapB), and M. sinensis ‘November Sunset’ (MapC), are typically winter hardy in Urbana. Nearly all MapA progeny plants survived and grew vigorously in spring 2014, whereas in MapB and MapC, many progeny plants did not survive the winter, and most of the survivors were severely damaged, with poor vigor. Negative correlations between overwintering ability and spring regrowth date and autumn dormancy date suggested that the genotypes most likely to survive winters were those that emerged early in spring and/or went dormant early in autumn. Using joint‐population analysis, we identified 53 quantitative trait loci (QTLs) for nine adaptation traits, including nine QTLs for overwintering ability and 11 for spring hardiness scores. Many biologically intuitive candidate genes were observed within or near the QTLs detected in this study, suggesting their validity and potential for further study.     

Identification of QTLs for hybrid fertility in inter-subspecific crosses of rice (Oryza sativa L.)

Two subspecies in rice, japonica and indica, have their own ecotypic traits. However, reproductive barriers such as spikelet sterility in hybrid progenies between subspecies have been an obstacle in breeding programs for combining desirable traits from both subspecies through inter-subspecific hybridization. The 166 F₉ RILs and two BC₁F₁s’ were analyzed for spikelet and pollen fertility with the parents and F₁ between Dasanbyeo (DS, indica) / TR22183 (TR, japonica). A frame map was constructed using a total of 218 polymorphic STS and SSR markers. In both BC₁F₁s’ of DS//DS/TR and TR//DS/TR, clusters of significant QTLs for spikelet and pollen fertility were identified on the short arm of chromosome 5 and chromosome 8. Nine and ten digenic epistatic interactions for DS//DS/TR and TR//DS//TR were identified, respectively. HF-QTLs were detected at the similar position with subspecies-specific markers and segregation distortion loci, implying that HF-QTLs might be associated with the differentiation of indica and japonica. Hybrid fertility/sterility and its relationship with other traits are discussed in relation to the reproductive barriers between subspecies of rice.     

Effect of high temperature and water stress on pollen germination and spikelet fertility in rice

In future climates, rice could more frequently be subjected to simultaneous high temperature and water stress during sensitive developmental stages such as flowering. In this study, five rice genotypes were exposed to high temperature, water stress and combined high temperature and water stress during flowering to quantify their response through spikelet fertility. Microscopic analyses revealed significant differences in anther dehiscence between treatments and genotypes, with a moderately high association with the number of germinated pollen grains on the stigma. There was a strong relationship between spikelet fertility and the number of germinated pollen on stigmas. Although, all three stress treatments resulted in spikelet sterility, high-temperature stress caused the highest sterility in all five genotypes. A cumulative linear decline in spikelet fertility with increasing duration of independent high-temperature stress and in combination with water stress was quantified. Better anther dehiscence, higher in vivo pollen germination, and higher spikelet fertility were observed in both the N22 accessions compared with IR64, Apo and Moroberekan under high temperature, water stress and combined stress, indicating its ability to tolerate multiple abiotic stresses.     

Assessing hybrid sterility in <Emphasis Type="Italic">Oryza glaberrima</Emphasis> × <Emphasis Type="Italic">O. sativa</Emphasis> hybrid progenies by PCR marker analysis and crossing with wide compatibility varieties

Interspecific crossing of the African indigenous rice Oryza glaberrima with Oryza sativa cultivars is hindered by crossing barriers causing 100% spikelet sterility in F₁ hybrids. Since hybrids are partially female fertile, fertility can be restored by back crossing (BC) to a recurrent male parent. Distinct genetic models on spikelet sterility have been developed predicting, e.g., the existence of a gamete eliminator and/or a pollen killer. Linkage of sterility to the waxy starch synthase gene and the chromogen gene C, both located on chromosome 6, have been demonstrated. We selected a segregating BC₂F₃ population of semi-sterile O. glaberrima × O. sativa indica hybrid progenies for analyses with PCR markers located at the respective chromosome-6 region. These analyses revealed that semi-sterile plants were heterozygous for a marker (OSR25) located in the waxy promoter, whereas fertile progenies were homozygous for the O. glaberrima allele. Adjacent markers showed no linkage to spikelet sterility. Semi-sterility of hybrid progenies was maintained at least until the F₄ progeny generation, suggesting the existence of a pollen killer in this plant material. Monitoring of reproductive plant development showed that spikelet sterility was at least partially due to an arrest of pollen development at the microspore stage. In order to address the question whether genes responsible for F₁ sterility in intraspecific hybrids (O. sativa indica × japonica) also cause spikelet sterility in interspecific hybrids, crossings with wide compatibility varieties (WCV) were performed. WCV accessions possess "neutral" S-loci (Sⁿ) improving fertility in intraspecific hybrids. This experiment showed that the tested Sⁿ-loci had no fertility restoring effect in F₁ interspecific hybrids. Pollen development was completely arrested at the microspore stage and grains were never obtained after selfing. This suggests that distinct or additional S-loci are responsible for sterility of O. glaberrima × O. sativa hybrids.Communicated by H.C. Becker     

TRANSCELLULAR BIOSYNTHESIS OF CYSTEINYL LEUKOTRIENES BY KUPFFER CELL—HEPATOCYTE COOPERATION IN RAT LIVER

We previously proposed that an enzymatic cooperation between Kupffer cells and hepatocytes may play an important role in cysteinyl leukotriene (LT) production in rat liver. Anin vitrotranscellular synthesis cysteinyl LTs by a Kupffer cell—hepatocyte coculture system was characterized here. Kupffer cells alone, with A23187 stimulation, did not generate cysteinyl LTs until supplemented either with isolated hepatocytes or with LTC₄synthase and glutathione, indicating that Kupffer cells can synthesize LTA₄but not convert it into LTC₄. In contrast, hepatocytes converted the LTA₄into cysteinyl LTs and further degraded the cysteinyl LTs. Cysteinyl LT production by the Kupffer cell—hapatocyte coculture system was optimized by addition of 1–3% serum albumin to the culture and by bringing the cell—cell distance closer to less than 3μ. Tumour necrosis factor also stimulated cysteinyl LT production by the coculture system. From these results, it is expected that the Kupffer cell—hepatocyte transcellular system for cysteinyl LT production actually functionsin vivo.