Left dorsolateral prefrontal cortex supports context-dependent prioritisation of off-task thought.

When environments lack compelling goals, humans often let their minds wander to thoughts with greater personal relevance; however, we currently do not understand how this context-dependent prioritisation process operates. Dorsolateral prefrontal cortex (DLPFC) maintains goal representations in a context-dependent manner. Here, we show this region is involved in prioritising off-task thought in an analogous way. In a whole brain analysis we established that neural activity in DLPFC is high both when 'on-task' under demanding conditions and 'off-task' in a non-demanding task. Furthermore, individuals who increase off-task thought when external demands decrease, show lower correlation between neural signals linked to external tasks and lateral regions of the DMN within DLPFC, as well as less cortical grey matter in regions sensitive to these external task relevant signals. We conclude humans prioritise daydreaming when environmental demands decrease by aligning cognition with their personal goals using DLPFC.

The role of energy intake and energy misreporting in the associations between eating patterns and adiposity.

BACKGROUND/OBJECTIVES: Research examining associations between eating occasion (EO) frequency and adiposity is inconclusive; studies examining the impact of energy misreporting are rare. This study examined associations between eating patterns and adiposity, with adjustment for energy misreporting, in a nationally representative sample of Australian adults. SUBJECTS/METHODS: Dietary intake was assessed via two 24-h recalls collected during the 2011-12 National Nutrition and Physical Activity Survey (n=4050 adults, aged ⩾19 years). Frequencies of all EOs, meals and snacks were calculated. Height, weight and waist circumference (WC) were measured. Energy misreporting was assessed as the ratio of energy intake to predicted energy expenditure (EI:EE). Energy misreporters were identified by EI:EE ratios, <0.68 or >1.32. Multivariate regression models assessed associations between eating patterns and body mass index (BMI), WC, overweight/obesity (BMI ⩾25 kg m-2) and central overweight/obesity (WC ⩾94 cm in men and ⩾80 cm in women). RESULTS: After adjustment for covariates and EI:EE, frequency of all EOs, meals (women only) and snacks was positively associated with WC and BMI (all P<0.01). Snack, but not meal frequency, was also associated with overweight/obesity (men: OR=1.22, 95% CI 1.07-1.39; women: OR=1.26, 95% CI 1.10-1.43) and central overweight/obesity (men: OR=1.17, 95% CI 1.04-1.32; women: OR=1.21, 95% CI 1.06-1.37). Multivariate analysis that excluded energy misreporters and adjusted for EI yielded either null or inverse associations (P<0.05). CONCLUSIONS: These findings suggest that the associations between eating patterns and adiposity are complicated by the role of EI and energy misreporting. Longitudinal research that considers the impact of EI and energy misreporting is needed to better understand the relationship between eating patterns and obesity.

Clustering of diet, physical activity and sedentary behaviour among Australian children: cross-sectional and longitudinal associations with overweight and obesity.

BACKGROUND/OBJECTIVES: Evidence suggests diet, physical activity (PA) and sedentary behaviour cluster together in children, but research supporting an association with overweight/obesity is equivocal. Furthermore, the stability of clusters over time is unknown. The aim of this study was to examine the clustering of diet, PA and sedentary behaviour in Australian children and cross-sectional and longitudinal associations with overweight/obesity. Stability of obesity-related clusters over 3 years was also examined. SUBJECTS/METHODS: Data were drawn from the baseline (T1: 2002/2003) and follow-up waves (T2: 2005/2006) of the Health Eating and Play Study. Parents of Australian children aged 5-6 (n=87) and 10-12 years (n=123) completed questionnaires. Children wore accelerometers and height and weight were measured. Obesity-related clusters were determined using K-medians cluster analysis. Multivariate regression models assessed cross-sectional and longitudinal associations between cluster membership, and body mass index (BMI) Z-score and weight status. Kappa statistics assessed cluster stability over time. RESULTS: Three clusters, labelled 'most healthy', 'energy-dense (ED) consumers who watch TV' and 'high sedentary behaviour/low moderate-to-vigorous PA' were identified at baseline and at follow-up. No cross-sectional associations were found between cluster membership, and BMI Z-score or weight status at baseline. Longitudinally, children in the 'ED consumers who watch TV' cluster had a higher odds of being overweight/obese at follow-up (odds ratio=2.8; 95% confidence interval: 1.1, 6.9; P<0.05). Tracking of cluster membership was fair to moderate in younger (K=0.24; P=0.0001) and older children (K=0.46; P<0.0001). CONCLUSIONS: This study identified an unhealthy cluster of TV viewing with ED food/drink consumption, which predicted overweight/obesity in a small longitudinal sample of Australian children. Cluster stability was fair to moderate over 3 years and is a novel finding. Prospective research in larger samples is needed to examine how obesity-related clusters track over time and influence the development of overweight and obesity.

Clustering of children's obesity-related behaviours: associations with sociodemographic indicators.

BACKGROUND/OBJECTIVES: Research suggests obesity-related behaviours cluster together in children and adolescents, but how these cluster patterns differ by sociodemographic indicators remains unclear. Furthermore, few studies examining clustering of behaviours have included younger children or an objective measure of physical activity (PA) and sedentary behaviour. Therefore, the aim of this study was to examine clustering patterns of diet, PA and sedentary behaviour in 5- to 6- and 10- to 12-year-old children, and their cross-sectional associations with sociodemographic indicators. SUBJECTS/METHODS: In this cross-sectional study, data from the baseline wave (2002/2003) of the Health Eating and Play study (HEAPS) were used. Questionnaires were completed by parents of Australian children aged 5-6 (n=362) and 10-12 years (n=610). Children wore accelerometers for up to 7 days. K-medians cluster analysis identified groups of children with similar diet, PA and sedentary behaviours. Chi-square tests assessed cluster differences by gender, maternal education and marital status. RESULTS: For each age group, three reliable and meaningful clusters were identified and labelled 'most healthy', 'energy-dense (ED) consumers who watch TV' and 'high sedentary behaviour/low moderate-to-vigorous PA (MVPA)'. Clusters varied by sociodemographic indicators. For example, a higher proportion of older girls comprised the 'high sedentary behaviour/low MVPA' cluster (χ(2)=22.4, P<0.001). Among both age groups, the 'ED consumers who watch TV' cluster comprised more children with lower educated mothers (younger children: χ(2)=34.9, P<0.001; older children: χ(2)=27.3, P<0.001). CONCLUSIONS: Identification of cluster patterns of obesity-related risk factors in children, and across sociodemographic groups may assist the targeting of public health initiatives, to those most in need.

Importance of sequences adjacent to the terminal tripeptide in the import of a peroxisomal Candida tropicalis protein in plant peroxisomes.

The peroxisome targeting signal (PTS) required for import of the rat acyl-CoA oxidase (AOX; EC 1.3.3.6) and the Candida tropicalis multifunctional protein (MFP) in plant peroxisomes was assessed in transgenic Arabidopsis thaliana (L.) Heynh. The native rat AOX accumulated in peroxisomes in A. thaliana cotyledons and targeting was dependent on the presence of the C-terminal tripeptide S-K-L. In contrast, the native C. tropicalis MFP, containing the consensus PTS sequence A-K-I was not targeted to plant peroxisomes. Modification of the carboxy terminus to the S-K-L tripeptide also failed to deliver the MFP to peroxisomes while addition of the last 34 amino acids of the Brassica napus isocitrate lyase, containing the terminal tripeptide S-R-M, enabled import of the fusion protein into peroxisomes. These results underline the influence of the amino acids adjacent to the terminal tripeptide of the C. tropicalis MFP on peroxisomal targeting, even in the context of a protein having a consensus PTS sequence S-K-L.

Chloroplast acclimation in leaves of Guzmania monostachia in response to high light.

Acclimation of leaves to high light (HL; 650 micromol m(-2) s(-1)) was investigated in the long-lived epiphytic bromeliad Guzmania monostachia and compared with plants maintained under low light (LL; 50 micromol m(-2) s(-1)). Despite a 60% decrease in total chlorophyll in HL-grown plants, the chlorophyll a/b ratio remained stable. Additionally, chloroplasts from HL-grown plants had a much lower thylakoid content and reduced granal stacking. Immunofluorescent labeling techniques were used to quantify the level of photosynthetic polypeptides. HL-grown plants had 30% to 40% of the content observed in LL-grown plants for the light-harvesting complex associated with photosystems I and II, the 33-kD photosystem II polypeptide, and Rubisco. These results were verified using conventional biochemical techniques, which revealed a comparable 60% decrease in Rubisco and total soluble protein. When expressed on a chlorophyll basis, the amount of protein and Rubisco was constant for HL- and LL-grown plants. Acclimation to HL involves a tightly coordinated adjustment of photosynthesis, indicating a highly regulated decrease in the number of photosynthetic units manifested at the level of the content of light-harvesting and electron transport components, the amount of Rubisco, and the induction of Crassulacean acid metabolism. This response occurs in mature leaves and may represent a strategy that is optimal for the resource-limited epiphytic niche.

The distinctive roles of five different ARC genes in the chloroplast division process in Arabidopsis.

ARC (accumulation and replication of chloroplasts) genes control different aspects of the chloroplast division process in higher plants. In order to establish the hierarchy of the ARC genes in the chloroplast division process and to provide evidence for their specific roles, double mutants were constructed between arc11, arc6, arc5, arc3 and arc1 in all combinations and phenotypically analysed. arc11 is a new nuclear recessive mutant with 29 chloroplasts compared with 120 in wild type. All the phenotypes of the double mutants are unambiguous. ARC1 down-regulates proplastid division but is on a separate pathway from ARC3, ARC5, ARC6 and ARC11. ARC6 initiates both proplastid and chloroplast division. ARC3 controls the rate of chloroplast expansion and ARC11 the central positioning of the final division plane in chloroplast division. ARC5 facilitates separation of the two daughter chloroplasts. ARC5 maps to chromosome 3 and ARC11 and ARC6 map approximately 60 cM apart on chromosome 5.

Synthesis of medium-chain-length polyhydroxyalkanoates in arabidopsis thaliana using intermediates of peroxisomal fatty acid beta-oxidation.

Polyhydroxyalkanoate (PHA) is a family of polymers composed primarily of R-3-hydroxyalkanoic acids. These polymers have properties of biodegradable thermoplastics and elastomers. Medium-chain-length PHAs (MCL-PHAs) are synthesized in bacteria by using intermediates of the beta-oxidation of alkanoic acids. To assess the feasibility of producing MCL-PHAs in plants, Arabidopsis thaliana was transformed with the PhaC1 synthase from Pseudomonas aeruginosa modified for peroxisome targeting by addition of the carboxyl 34 amino acids from the Brassica napus isocitrate lyase. Immunocytochemistry demonstrated that the modified PHA synthase was appropriately targeted to leaf-type peroxisomes in light-grown plants and glyoxysomes in dark-grown plants. Plants expressing the PHA synthase accumulated electron-lucent inclusions in the glyoxysomes and leaf-type peroxisomes, as well as in the vacuole. These inclusions were similar to bacterial PHA inclusions. Analysis of plant extracts by GC and mass spectrometry demonstrated the presence of MCL-PHA in transgenic plants to approximately 4 mg per g of dry weight. The plant PHA contained saturated and unsaturated 3-hydroxyalkanoic acids ranging from six to 16 carbons with 41% of the monomers being 3-hydroxyoctanoic acid and 3-hydroxyoctenoic acid. These results indicate that the beta-oxidation of plant fatty acids can generate a broad range of R-3-hydroxyacyl-CoA intermediates that can be used to synthesize MCL-PHAs.

Immunofluorescent quantitation of chloroplast proteins.

Using scanning light microscopy software to detect and measure immunofluorescence in leaf sections Rubisco concentration in situ in chloroplasts has been accurately determined throughout development. The fluorescence measurements were calibrated by comparison with values for Rubisco accumulation obtained from rocket immuno-electrophoresis profiles of soluble protein from isolated cells and from chloroplasts using a purified sample of Rubisco as the standard. It has been shown that in situ immunofluorescence can be used for cytoquantitation of proteins within individual chloroplasts to a sensitivity of 1fg and also for the comparison of the protein levels in adjacent chloroplasts and cells. Several important applications of this new technique are discussed.

Castor bean isocitrate lyase lacking the putative peroxisomal targeting signal 1 ARM is imported into plant peroxisomes both in vitro and in vivo.

To understand and manipulate plant peroxisomal protein targeting, it is important to establish the universality or otherwise of targeting signals. Contradictory results have been published concerning the nature and location of the glyoxysomal/peroxisomal targeting signal of isocitrate lyase (ICL). L.J. Olsen, W.F. Ettinger, B. Damsz, K. Matsudaira, A. Webb, and J.J. Harada ([1993] Plant Cell 5: 941-952) concluded that the last 5 amino acids (AKSRM) of Brassica napus ICL were sufficient and the last 37 amino acids were necessary for targeting to Arabidopsis leaf peroxisomes. In contrast, R. Behari and A. Baker ([1993]) J Biol Chem 268: 7315-7322) could find no requirement for the almost identical carboxy-terminal sequence AKARM for import of Ricinus communis ICL into isolated sunflower cotyledon glyoxysomes. To resolve this discrepancy, the import characteristics of a mutant R. communis ICL lacking the last 19 amino acids of the carboxy terminus was studied. ICL delta 19 was able to be imported by isolated sunflower glyoxysomes and by tobacco leaf peroxisomes when expressed transgenically. These results demonstrate that the in vitro import system faithfully reflects targeting in vivo, and that the source of the organelles (Arabidopsis versus sunflower, leaf peroxisomes versus seed glyoxysomes) is not responsible for observed differences between B. napus and R. communis ICL. The R. communis enzyme would therefore appear to possess an additional glyoxysome/peroxisome targeting signal that is lacking in the B. napus protein.

Characterization of chloroplast division using the Arabidopsis mutant arc5.

arc5 is a chloroplast division mutant of Arabidopsis thaliana. To identify the role of ARC5 in the chloroplast replication process we have followed the changes in arc5 chloroplasts during their perturbed division. ARC5 does not affect proplastid division but functions at a later stage in chloroplast development. Chloroplasts in developing mesophyll cells of arc5 leaves do not increase in number and all of the chloroplasts in mature leaf cells show a central constriction. Young arc5 chloroplasts are capable of initiating the division process but fail to complete daughter-plastid separation. Wild-type plastids increase in number to a mean of 121 after completing the division process, but in the mutant arc5 the approximately 13 plastids per cell are still centrally constricted but much enlarged. As the arc5 chloroplasts expand and elongate without dividing, the internal thylakoid membrane structure becomes flexed into an undulating ribbon. We conclude that the ARC5 gene is necessary for the completion of the last stage of chloroplast division when the narrow isthmus breaks, causing the separation of the daughter plastids.

A putative Mg chelatase subunit from Arabidopsis thaliana cv C24. Sequence and transcript analysis of the gene, import of the protein into chloroplasts, and in situ localization of the transcript and protein.

We have isolated and sequenced a cDNA from Arabidopsis thaliana cv C24 that encodes a putative Mg chelatase subunit. The deduced amino acid sequence shows a very high level of identity to a gene previously characterized from Antirrhinum majus (olive and also high similarity to bchH, a bacterial gene involved in the Mg chelatase reaction of bacteriochlorophyll biosynthesis. We suggest that this gene be called CHL H. Northern blot analyses were used to investigate the expression of CHL H, another putative Mg chelatase gene, ch-42, and ferrochelatase. The CHL H transcript was observed to undergo a dramatic diurnal variation, rising almost to its maximum level by the end of the dark period, then increasing slightly at the onset of the light and declining steadily to a minimum by the end of the light period; in contrast, transcripts for ch-42 and ferrochelatase remained constant. A model is proposed in which the CHL H protein plays a role in regulating the levels of chlorophyll during this cycle. In situ hybridization revealed that the transcripts are located over the surface of the chloroplasts, a feature in common with transcripts for the ch-42 gene. The CHL H protein was imported into the stromal compartment of the chloroplast and processed in an in vitro assay. Immunoblotting showed that the distribution of CHL H protein between the stroma and chloroplast membranes varies depending on the concentration of Mg+. In situ immunofluorescence was used to establish that the CHL H and CH-42 proteins are localized within the chloroplast in vivo.

Subcellular Visualization of Gene Transcripts Encoding Key Proteins of the Chlorophyll Accumulation Process in Developing Chloroplasts.

The coordination of the synthesis of chlorophyll (Chl) and light-harvesting Chl proteins was determined by observing the sequence of appearance of the specific mRNAs for the nuclear genes CHLH, Por, and Lhcb1*2 (AB180). CHLH encodes a magnesium protoporphyrin chelatase subunit that is involved in the first committed step in Chl biosynthesis; Por encodes protochlorophyllide oxidoreductase, which catalyzes the penultimate and only light-dependent step in Chl biosynthesis; and Lhcb1*2 encodes light-harvesting Chl a/b binding protein of the type-1 light-harvesting complex of photosystem II. Using digoxigenin-labeled antisense and sense RNA probes and a highly sensitive in situ hybridization technique, we have visualized the first appearance of the specific mRNAs in postmitotic mesophyll cells of developing 7-d-old wheat leaves (Triticum aestivum cv Maris dove). The transcripts for CHLH and POR are detectable in the youngest (18 h postmitotic) leaf tissue containing dividing cells; light-harvesting complex of photosystem II transcripts appear 12 h later. This is consistent with a requirement for accumulation of Chl before synthesis of Chl a/b binding protein can proceed at a high rate. All of the transcripts are most abundant in mesophyll cells. In the first leaf the POR message is initially restricted to the palisade, but 12 h later it is also present in the spongy mesophyll cells. All three transcripts aggregated around the surface of the chloroplasts, suggesting that translation may occur preferentially in the vicinity of the target organelle for the primary translation products.

arc6, an extreme chloroplast division mutant of Arabidopsis also alters proplastid proliferation and morphology in shoot and root apices.

The arc6 (accumulation and replication of chloroplasts) mutant of Arabidopsis has only two greatly enlarged chloroplasts per mature leaf mesophyll cell compared with ninety chloroplasts per cell in the wild type. The mutation is a single nuclear gene and the plant phenotype is normal. Shoot and root apical meristems of arc6 plants have been examined to determine how early during plastid development the mutant arc6 phenotype can be recognised. In the cells of the arc6 apical meristem there are only two proplastids, which are larger than wild type with a highly variable morphology. In the cells of the leaf primordia where differentiation of proplastids to chloroplasts occurs arc6 plastids are larger and at a more advanced developmental stage than wild-type plastids. In arc6 root cells statoliths and other plastids also show grossly abnormal morphology and the statoliths are greatly increased in size. During arc6 stomatal guard cell development the perturbation in proplastid population dynamics affects plastid segregation and 30% of stomata lack plastids in one or both guard cells. Our evidence would suggest that ARC6 is expressed throughout the vegetative cells of the Arabidopsis seedling with major effects on both the proplastid phenotype and the proplastid population. ARC6 is the first gene to be identified in Arabidopsis which has a global effect on plastid development in cells arising from both the shoot and root meristems, and is of major importance in the nuclear control of plastid differentiation in higher plants.

Mitochondria Increase Three-Fold and Mitochondrial Proteins and Lipid Change Dramatically in Postmeristematic Cells in Young Wheat Leaves Grown in Elevated CO2.

A dramatic stimulation in mitochondrial biogenesis during the very early stages of leaf development was observed in young wheat plants (Triticum aestivum cv Hereward) grown in elevated CO2 (650 [mu]L L-1). An almost 3-fold increase in the number of mitochondria was observed in the very young leaf cells at the base of the first leaf of a 7-d-old wheat plant. In the same cells large increases in the accumulation of a mitochondrial chaperonin protein and the mitochondrial 2-oxoglutarate dehydrogenase complex and pyruvate dehydrogenase complex were detected by immunolabeling. Furthermore, the basal segment also shows a large increase in the rate of radiolabeling of diphosphatidylglycerol, a lipid confined to the inner mitochondrial membrane. This dramatic response in very young leaf cells to elevated CO2 suggests that the numerous documented positive effects of elevated CO2 on wheat leaf development are initiated as early as 12 h postmitosis.

Significant Changes in Cell and Chloroplast Development in Young Wheat Leaves (Triticum aestivum cv Hereward) Grown in Elevated CO2.

Cell and chloroplast development were characterized in young Triticum aestivum cv Hereward leaves grown at ambient (350 [mu]L L-1) or at elevated (650 [mu]L L-1) CO2. In elevated CO2, cell and chloroplast expansion was accelerated by 10 and 25%, respectively, in the first leaf of 7-d-old wheat plants without disruption to the leaf developmental pattern. Elevated CO2 did not affect the number of chloroplasts in relation to mesophyll cell size or the linear relationship between chloroplast number or size and mesophyll cell size. No major changes in leaf anatomy or in chloroplast ultrastructure were detected as a result of growth in elevated CO2, but there was a marked reduction in starch accumulation. In leaf sections fluorescently tagged antisera were used to visualize and quantitate the amount of cytochrome f, the [alpha]- and [beta]-subunits of the coupling factor 1 in ATP synthase, D1 protein of the photosystem II reaction center, the 33-kD protein of the extrinsic oxygen-evolving complex, subunit II of photosystem I, and ribulose-1,5-bisphosphate carboxylase/oxygenase. A significant finding was that in 10 to 20% of the mesophyll cells grown in elevated CO2 the 33-kD protein of the extrinsic oxygen-evolving complex of photosystem II and cytochrome f were deficient by 75%, but the other proteins accumulated normally.

arc6, A Fertile Arabidopsis Mutant with Only Two Mesophyll Cell Chloroplasts.

A novel mutant of Arabidopsis thaliana, arc6 (accumulation and replication of chloroplasts), has been isolated from a transfer DNA-mutagenized population of Arabidopsis seedlings. arc6 has the most extreme arc mutant phenotype we have yet described, with only one to three chloroplasts per leaf mesophyll cell compared to a mean of 83 in cells of the wild-type var Wassilewskija. The chloroplasts of arc6 are 20-fold larger than wild-type chloroplasts.Chloroplast division is almost certainly precluded in arc6 mesophyll cells, since chloroplast number per cell does not increase during mesophyll cell expansion. arc6 chloroplasts are long and thin in cross-section and only one-half the width of wild-type chloroplasts and the arrangement of thylakoid membranes is largely unaltered. arc6 segregates as a monogenic recessive nuclear mutation in a normal Mendelian manner and the arc6 phenotype is stably inherited for at least four generations. arc6 plants grow normally and are fertile, although the rosette leaves appear curled and twisted. arc6 plants accumulate 70 to 75% of the biomass of wild type. The phenotype of this novel mutant is discussed in relation to the nature of the control of chloroplast division in leaf cells.

A Genetic Analysis of Chloroplast Division and Expansion in Arabidopsis thaliana.

A nuclear recessive mutant of Arabidopsis thaliana, arc5, has been isolated in which there is no significant increase in chloroplast number during leaf mesophyll cell expansion and in which there are only 13 chloroplasts per mesophyll cell compared with 121 in wild-type cells. Mature arc5 chloroplasts in fully expanded mesophyll cells are 6-fold larger than in wild-type cells. A large proportion of arc5 chloroplasts also show some degree of central constriction, suggesting that the mutation has prevented the completion of the chloroplast division process. To examine the interaction of arc loci, a double mutant was constructed between arc1, a mutant possessing many small chloroplasts, and arc5. A second double mutant was also constructed between arc3, a previously discovered mutant also possessing few large chloroplasts per cell, and arc1. Analysis of these double mutants shows that chloroplast number per mesophyll cell is greater when arc5 and arc3 mutations are expressed in the arc1 background than when expressed alone. The cell-specific nature of arc mutants was also analyzed. The phenotypic traits characteristic of arc3 and arc5 are a reduction in chloroplast number and an increase in chloroplast size in mesophyll cells: these changes are also observed in reduced form in the epidermal and guard cell chloroplasts of arc3 and arc5 plants. Analysis of parenchyma sheath cell chloroplasts suggests that in leaves of arc1 plants the normal developmental distinction between mesophyll and parenchyma sheath chloroplasts is perturbed. The relevance of these findings to the analysis of the control of chloroplast division in mesophyll cells is discussed.

Recognition of Peroxisomes by Immunofluorescence in Transformed and Untransformed Tobacco Cells.

We report the visualization of peroxisomes in tobacco (Nicotiana tabacum) leaves using fluorescently labeled antibodies to glycolate oxidase. In transgenic tobacco leaves the expression of isocitrate lyase was also visualized. In dual probing experiments both enzymes were shown to be present together in all peroxisomes in transgenic tobacco leaves.

Chloroplast Division and Expansion Is Radically Altered by Nuclear Mutations in Arabidopsis thaliana.

We have isolated three mutants of Arabidopsis thaliana in which there is a sevenfold change in chloroplast number in fully expanded leaf mesophyll cells and increases and decreases in chloroplast number are compensated for by changes in chloroplast size. The changes are stably inherited in sexual crosses for three generations and mutant phenotypes are effected by changes at single recessive nuclear loci, termed arc loci. This is the first report of large, stably inherited changes in chloroplast number in higher plants, and represents a major advance toward the genetic dissection of the control of chloroplast division.