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Immunodeficiency, Centromeric instability and Facial anomalies (ICF) syndrome is a rare genetic disorder characterized by variable immunodeficiency. More than half of the affected individuals show mild to severe intellectual disability at early onset. This disorder is genetically heterogeneous and ZBTB24 is the causative gene of the subtype 2, accounting for about 30% of the ICF cases. ZBTB24 is a multifaceted transcription factor belonging to the Zinc-finger and BTB domain-containing protein family, which are key regulators of developmental processes. Aberrant DNA methylation is the main molecular hallmark of ICF syndrome. The functional link between ZBTB24 deficiency and DNA methylation errors is still elusive. Here, we generated a novel ICF2 disease model by deriving induced pluripotent stem cells (iPSCs) from peripheral CD34+-blood cells of a patient homozygous for the p.Cys408Gly mutation, the most frequent missense mutation in ICF2 patients and which is associated with a broad clinical spectrum. The mutation affects a conserved cysteine of the ZBTB24 zinc-finger domain, perturbing its function as transcriptional activator. ICF2-iPSCs recapitulate the methylation defects associated with ZBTB24 deficiency, including centromeric hypomethylation. We validated that the mutated ZBTB24 protein loses its ability to directly activate expression of CDCA7 and other target genes in the patient-derived iPSCs. Upon hematopoietic differentiation, ICF2-iPSCs showed decreased vitality and a lower percentage of CD34+/CD43+/CD45+ progenitors. Overall, the ICF2-iPSC model is highly relevant to explore the role of ZBTB24 in DNA methylation homeostasis and provides a tool to investigate the early molecular events linking ZBTB24 deficiency to the ICF2 clinical phenotype.
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Células Madre Pluripotentes Inducidas , Fenotipo , Enfermedades de Inmunodeficiencia Primaria , Proteínas Represoras , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Enfermedades de Inmunodeficiencia Primaria/genética , Proteínas Represoras/genética , Proteínas Represoras/deficiencia , Metilación de ADN , Síndromes de Inmunodeficiencia/genética , Masculino , Mutación , Femenino , Cara/anomalías , Proteínas NuclearesRESUMEN
Oocyte maturation and preimplantation embryo development are critical to successful pregnancy outcomes and the correct establishment and maintenance of genomic imprinting. Thanks to novel technologies and omics studies in human patients and mouse models, the importance of the proteins associated with the cytoplasmic lattices (CPLs), highly abundant structures found in the cytoplasm of mammalian oocytes and preimplantation embryos, in the maternal to zygotic transition is becoming increasingly evident. This review highlights the recent discoveries on the role of these proteins in protein storage and other oocyte cytoplasmic processes, epigenetic reprogramming, and zygotic genome activation (ZGA). A better comprehension of these events may significantly improve clinical diagnosis and pave the way for targeted interventions aiming to correct or mitigate female fertility issues and genomic imprinting disorders.
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Maternal inactivation of genes encoding components of the subcortical maternal complex (SCMC) and its associated member, PADI6, generally results in early embryo lethality. In humans, SCMC gene variants were found in the healthy mothers of children affected by multilocus imprinting disturbances (MLID). However, how the SCMC controls the DNA methylation required to regulate imprinting remains poorly defined. We generated a mouse line carrying a Padi6 missense variant that was identified in a family with Beckwith-Wiedemann syndrome and MLID. If homozygous in female mice, this variant resulted in interruption of embryo development at the two-cell stage. Single-cell multiomic analyses demonstrated defective maturation of Padi6 mutant oocytes and incomplete DNA demethylation, down-regulation of zygotic genome activation (ZGA) genes, up-regulation of maternal decay genes, and developmental delay in two-cell embryos developing from Padi6 mutant oocytes but little effect on genomic imprinting. Western blotting and immunofluorescence analyses showed reduced levels of UHRF1 in oocytes and abnormal localization of DNMT1 and UHRF1 in both oocytes and zygotes. Treatment with 5-azacytidine reverted DNA hypermethylation but did not rescue the developmental arrest of mutant embryos. Taken together, this study demonstrates that PADI6 controls both nuclear and cytoplasmic oocyte processes that are necessary for preimplantation epigenetic reprogramming and ZGA.
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Oocitos , Cigoto , Animales , Niño , Femenino , Humanos , Ratones , Proteínas Potenciadoras de Unión a CCAAT/genética , Citoplasma/genética , Citoplasma/metabolismo , Metilación de ADN/genética , Desarrollo Embrionario/genética , Impresión Genómica/genética , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
Initially described as a triad of immunodeficiency, congenital heart defects and hypoparathyroidism, 22q11.2 deletion syndrome (22q11.2DS) now encompasses a great amount of abnormalities involving different systems. Approximately 85% of patients share a 3 Mb 22q11.2 region of hemizygous deletion in which 46 protein-coding genes are included. However, the hemizygosity of the genes of this region cannot fully explain the clinical phenotype and the phenotypic variability observed among patients. Additional mutations in genes located outside the deleted region, leading to "dual diagnosis", have been described in 1% of patients. In some cases, the hemizygosity of the 22q11.2 region unmasks autosomal recessive conditions due to additional mutations on the non-deleted allele. Some of the deleted genes play a crucial role in gene expression regulation pathways, involving the whole genome. Typical miRNA expression patterns have been identified in 22q11.2DS, due to an alteration in miRNA biogenesis, affecting the expression of several target genes. Also, a methylation epi-signature in CpG islands differentiating patients from controls has been defined. Herein, we summarize the evidence on the genetic and epigenetic mechanisms implicated in the pathogenesis of the clinical manifestations of 22q11.2 DS. The review of the literature confirms the hypothesis that the 22q11.2DS phenotype results from a network of interactions between deleted protein-coding genes and altered epigenetic regulation.
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Síndrome de DiGeorge , Cardiopatías Congénitas , MicroARNs , Humanos , Síndrome de DiGeorge/genética , Epigénesis Genética , Fenotipo , Cardiopatías Congénitas/genéticaRESUMEN
Parent-of-origin-dependent gene expression of a few hundred human genes is achieved by differential DNA methylation of both parental alleles. This imprinting is required for normal development, and defects in this process lead to human disease. Induced pluripotent stem cells (iPSCs) serve as a valuable tool for in vitro disease modeling. However, a wave of de novo DNA methylation during reprogramming of iPSCs affects DNA methylation, thus limiting their use. The DNA methyltransferase 3B (DNMT3B) gene is highly expressed in human iPSCs; however, whether the hypermethylation of imprinted loci depends on DNMT3B activity has been poorly investigated. To explore the role of DNMT3B in mediating de novo DNA methylation at imprinted DMRs, we utilized iPSCs generated from patients with immunodeficiency, centromeric instability, facial anomalies type I (ICF1) syndrome that harbor biallelic hypomorphic DNMT3B mutations. Using a whole-genome array-based approach, we observed a gain of methylation at several imprinted loci in control iPSCs but not in ICF1 iPSCs compared to their parental fibroblasts. Moreover, in corrected ICF1 iPSCs, which restore DNMT3B enzymatic activity, imprinted DMRs did not acquire control DNA methylation levels, in contrast to the majority of the hypomethylated CpGs in the genome that were rescued in the corrected iPSC clones. Overall, our study indicates that DNMT3B is responsible for de novo methylation of a subset of imprinted DMRs during iPSC reprogramming and suggests that imprinting is unstable during a specific time window of this process, after which the epigenetic state at these regions becomes resistant to perturbation.
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Síndromes de Inmunodeficiencia , Células Madre Pluripotentes Inducidas , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , ADN (Citosina-5-)-Metiltransferasas/genética , ADN (Citosina-5-)-Metiltransferasas/metabolismo , Metilación de ADN , Mutación , Síndromes de Inmunodeficiencia/genética , Impresión GenómicaRESUMEN
Imprinting disorders are congenital diseases caused by dysregulation of genomic imprinting, affecting growth, neurocognitive development, metabolism and cancer predisposition. Overlapping clinical features are often observed among this group of diseases. In rare cases, two fully expressed imprinting disorders may coexist in the same patient. A dozen cases of this type have been reported so far. Most of them are represented by individuals affected by Beckwith-Wiedemann spectrum (BWSp) and Transient Neonatal Diabetes Mellitus (TNDM) or BWSp and Pseudo-hypoparathyroidism type 1B (PHP1B). All these patients displayed Multilocus imprinting disturbances (MLID). Here, we report the first case of co-occurrence of BWS and PHP1B in the same individual in absence of MLID. Genome-wide methylation and SNP-array analyses demonstrated loss of methylation of the KCNQ1OT1:TSS-DMR on chromosome 11p15.5 as molecular cause of BWSp, and upd(20)pat as cause of PHP1B. The absence of MLID and the heterodisomy of chromosome 20 suggests that BWSp and PHP1B arose through distinct and independent mechanism in our patient. However, we cannot exclude that the rare combination of the epigenetic defect on chromosome 11 and the UPD on chromosome 20 may originate from a common so far undetermined predisposing molecular lesion. A better comprehension of the molecular mechanisms underlying the co-occurrence of two imprinting disorders will improve genetic counselling and estimate of familial recurrence risk of these rare cases. Furthermore, our study also supports the importance of multilocus molecular testing for revealing MLID as well as complex cases of imprinting disorders.
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CRC is an adult-onset carcinoma representing the third most common cancer and the second leading cause of cancer-related deaths in the world. EO-CRC (<45 years of age) accounts for 5% of the CRC cases and is associated with cancer-predisposing genetic factors in half of them. Here, we describe the case of a woman affected by BWSp who developed EO-CRC at age 27. To look for a possible molecular link between BWSp and EO-CRC, we analysed her whole-genome genetic and epigenetic profiles in blood, and peri-neoplastic and neoplastic colon tissues. The results revealed a general instability of the tumor genome, including copy number and methylation changes affecting genes of the WNT signaling pathway, CRC biomarkers and imprinted loci. At the germline level, two missense mutations predicted to be likely pathogenic were found in compound heterozygosity affecting the Cystic Fibrosis (CF) gene CFTR that has been recently classified as a tumor suppressor gene, whose dysregulation represents a severe risk factor for developing CRC. We also detected constitutional loss of methylation of the KCNQ1OT1:TSS-DMR that leads to bi-allelic expression of the lncRNA KCNQ1OT1 and BWSp. Our results support the hypothesis that the inherited CFTR mutations, together with constitutional loss of methylation of the KCNQ1OT1:TSS-DMR, initiate the tumorigenesis process. Further somatic genetic and epigenetic changes enhancing the activation of the WNT/beta-catenin pathway likely contributed to increase the growth advantage of cancer cells. Although this study does not provide any conclusive cause-effect relationship between BWSp and CRC, it is tempting to speculate that the imprinting defect of BWSp might accelerate tumorigenesis in adult cancer in the presence of predisposing genetic variants.
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Silver-Russell syndrome is an imprinting disorder characterised by pre- and post-natal growth retardation and several heterogeneous molecular defects affecting different human genomic loci. In the majority of cases, the molecular defect is the loss of methylation (LOM) of the H19/IGF2 differentially methylated region (DMR, also known as IC1) at the telomeric domain of the 11p15.5 imprinted genes cluster, which causes the altered expression of the growth controlling genes, IGF2 and H19. Very rarely, the LOM also affects the KCNQ1OT1 DMR (also known as IC2) at the centromeric domain, resulting in an SRS phenotype by an unknown mechanism. In this study, we report on two cases with SRS features and a LOM of either IC1 and IC2. In one case, this rare and complex epimutation was secondary to a de novo mosaic in cis maternal duplication, involving the entire telomeric 11p15.5 domain and part of the centromeric domain but lacking CDKN1C. In the second case, neither the no 11p15.5 copy number variant nor the maternal-effect subcortical maternal complex (SCMC) variant were found to be associated with the epimutation, suggesting that it arose as a primary event. Our findings further add to the complexity of the molecular genetics of SRS and indicate how the LOM in both 11p15.5 DMRs may result from different molecular mechanisms.
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Síndrome de Silver-Russell , Humanos , Síndrome de Silver-Russell/genética , Impresión Genómica , Metilación de ADN/genética , Fenotipo , Variaciones en el Número de Copia de ADNRESUMEN
BACKGROUND: Imprinting Control Regions (ICRs) are CpG-rich sequences acquiring differential methylation in the female and male germline and maintaining it in a parental origin-specific manner in somatic cells. Despite their expected high mutation rate due to spontaneous deamination of methylated cytosines, ICRs show conservation of CpG-richness and CpG-containing transcription factor binding sites in mammalian species. However, little is known about the mechanisms contributing to the maintenance of a high density of methyl CpGs at these loci. RESULTS: To gain functional insights into the mechanisms for maintaining CpG methylation, we sought to identify the proteins binding the methylated allele of the ICRs by determining the interactors of ZFP57 that recognizes a methylated hexanucleotide motif of these DNA regions in mouse ESCs. By using a tagged approach coupled to LC-MS/MS analysis, we identified several proteins, including factors involved in mRNA processing/splicing, chromosome organization, transcription and DNA repair processes. The presence of the post-replicative mismatch-repair (MMR) complex components MSH2 and MSH6 among the identified ZFP57 interactors prompted us to investigate their DNA binding profile by chromatin immunoprecipitation and sequencing. We demonstrated that MSH2 was enriched at gene promoters overlapping unmethylated CpG islands and at repeats. We also found that both MSH2 and MSH6 interacted with the methylated allele of the ICRs, where their binding to DNA was mediated by the ZFP57/KAP1 complex. CONCLUSIONS: Our findings show that the MMR complex is concentrated on gene promoters and repeats in mouse ESCs, suggesting that maintaining the integrity of these regions is a primary function of highly proliferating cells. Furthermore, the demonstration that MSH2/MSH6 are recruited to the methylated allele of the ICRs through interaction with ZFP57/KAP1 suggests a role of the MMR complex in the maintenance of the integrity of these regulatory regions and evolution of genomic imprinting in mammalian species.
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Metilación de ADN , Proteínas Represoras , Animales , Cromatografía Liquida , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Femenino , Impresión Genómica , Mamíferos/metabolismo , Ratones , Proteína 2 Homóloga a MutS/genética , Proteína 2 Homóloga a MutS/metabolismo , Proteínas Represoras/metabolismo , Espectrometría de Masas en TándemRESUMEN
BACKGROUND: Beckwith-Wiedemann syndrome (BWS) and Pseudohypoparathyroidism type 1B (PHP1B) are imprinting disorders (ID) caused by deregulation of the imprinted gene clusters located at 11p15.5 and 20q13.32, respectively. In both of these diseases a subset of the patients is affected by multi-locus imprinting disturbances (MLID). In several families, MLID is associated with damaging variants of maternal-effect genes encoding protein components of the subcortical maternal complex (SCMC). However, frequency, penetrance and recurrence risks of these variants are still undefined. In this study, we screened two cohorts of BWS patients and one cohort of PHP1B patients for the presence of MLID, and analysed the positive cases for the presence of maternal variants in the SCMC genes by whole exome-sequencing and in silico functional studies. RESULTS: We identified 10 new cases of MLID associated with the clinical features of either BWS or PHP1B, in which segregate 13 maternal putatively damaging missense variants of the SCMC genes. The affected genes also included KHDC3L that has not been associated with MLID to date. Moreover, we highlight the possible relevance of relatively common variants in the aetiology of MLID. CONCLUSION: Our data further add to the list of the SCMC components and maternal variants that are involved in MLID, as well as of the associated clinical phenotypes. Also, we propose that in addition to rare variants, common variants may play a role in the aetiology of MLID and imprinting disorders by exerting an additive effect in combination with rarer putatively damaging variants. These findings provide useful information for the molecular diagnosis and recurrence risk evaluation of MLID-associated IDs in genetic counselling.
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Síndrome de Beckwith-Wiedemann , Seudohipoparatiroidismo , Síndrome de Beckwith-Wiedemann/diagnóstico , Síndrome de Beckwith-Wiedemann/genética , Metilación de ADN , Impresión Genómica , Humanos , Proteínas/genética , Seudohipoparatiroidismo/genética , SeudohipoparatiroidismoRESUMEN
ZFP57 is required to maintain the germline-marked differential methylation at imprinting control regions (ICRs) in mouse embryonic stem cells (ESCs). Although DNA methylation has a key role in genomic imprinting, several imprinted genes are controlled by different mechanisms, and a comprehensive study of the relationship between DMR methylation and imprinted gene expression is lacking. To address the latter issue, we differentiated wild-type and Zfp57-/- hybrid mouse ESCs into neural precursor cells (NPCs) and evaluated allelic expression of imprinted genes. In mutant NPCs, we observed a reduction of allelic bias of all the 32 genes that were imprinted in wild-type cells, demonstrating that ZFP57-dependent methylation is required for maintaining or acquiring imprinted gene expression during differentiation. Analysis of expression levels showed that imprinted genes expressed from the non-methylated chromosome were generally up-regulated, and those expressed from the methylated chromosome were down-regulated in mutant cells. However, expression levels of several imprinted genes acquiring biallelic expression were not affected, suggesting the existence of compensatory mechanisms that control their RNA level. Since neural differentiation was partially impaired in Zfp57-mutant cells, this study also indicates that imprinted genes and/or non-imprinted ZFP57-target genes are required for proper neurogenesis in cultured ESCs.
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Metilación de ADN/genética , Impresión Genómica/genética , Células Madre Embrionarias de Ratones/metabolismo , Proteínas Represoras/genética , Animales , Diferenciación Celular/genética , Cromosomas/genética , Regulación del Desarrollo de la Expresión Génica/genética , Ratones , Células-Madre Neurales/metabolismoRESUMEN
The reciprocal parent of origin-specific expression of H19 and IGF2 is controlled by the H19/IGF2:IG-DMR (IC1), whose maternal allele is unmethylated and acts as a CTCF-dependent insulator. In humans, internal IC1 deletions are associated with Beckwith-Wiedemann syndrome (BWS) and Silver-Russell syndrome (SRS), depending on their parental origin. These genetic mutations result in aberrant DNA methylation, deregulation of IGF2/H19 and disease with incomplete penetrance. However, the mechanism linking the microdeletions to altered molecular and clinical phenotypes remains unclear. To address this issue, we have previously generated and characterized two knock-in mouse lines with the human wild-type (hIC1wt) or mutant (hIC1∆2.2) IC1 allele replacing the endogenous mouse IC1 (mIC1). Here, we report an additional knock-in line carrying a mutant hIC1 allele with an internal 1.8 kb deletion (hIC1∆1.8). The phenotype of these mice is different from that of the hIC1∆2.2-carrying mice, partially resembling hIC1wt animals. Indeed, proper H19 and Igf2 imprinting and normal growth phenotype were evident in the mice with maternal transmission of hIC1Δ1.8, while low DNA methylation and non-viable phenotype characterize its paternal transmission. In contrast to hIC1wt, E15.5 embryos that paternally inherit hIC1Δ1.8 displayed variegated hIC1 methylation. In addition, increased Igf2 expression, correlating with increased body weight, was found in one third of these mice. Chromatin immunoprecipitation experiments in mouse embryonic stem cells carrying the three different hIC1 alleles demonstrate that the number of CTCF target sites influences its binding to hIC1, indicating that in the mouse, CTCF binding is key to determining hIC1 methylation and Igf2 expression.
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Síndrome de Beckwith-Wiedemann , ARN Largo no Codificante , Animales , Síndrome de Beckwith-Wiedemann/genética , Sitios de Unión , Factor de Unión a CCCTC/genética , Metilación de ADN/genética , Impresión Genómica , Factor II del Crecimiento Similar a la Insulina/genética , Factor II del Crecimiento Similar a la Insulina/metabolismo , Ratones , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismoRESUMEN
ZBTB2 is a protein belonging to the BTB/POZ zinc-finger family whose members typically contain a BTB/POZ domain at the N-terminus and several zinc-finger domains at the C-terminus. Studies have been carried out to disclose the role of ZBTB2 in cell proliferation, in human cancers and in regulating DNA methylation. Moreover, ZBTB2 has been also described as an ARF, p53 and p21 gene repressor as well as an activator of genes modulating pluripotency. In this scenario, ZBTB2 seems to play many functions likely associated with other proteins. Here we report a picture of the ZBTB2 protein partners in U87MG cell line, identified by high-resolution mass spectrometry (MS) that highlights the interplay between ZBTB2 and chromatin remodeling multiprotein complexes. In particular, our analysis reveals the presence, as ZBTB2 candidate interactors, of SMARCA5 and BAZ1B components of the chromatin remodeling complex WICH and PBRM1, a subunit of the SWI/SNF complex. Intriguingly, we identified all the subunits of the NuRD complex among the ZBTB2 interactors. By co-immunoprecipitation experiments and ChIP-seq analysis we definitely identify ZBTB2 as a new partner of the NuRD complex.
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Complejo Desacetilasa y Remodelación del Nucleosoma Mi-2/metabolismo , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Adenosina Trifosfatasas/metabolismo , Línea Celular Tumoral , Cromatina/genética , Proteínas Cromosómicas no Histona/metabolismo , ADN/genética , Metilación de ADN , Proteínas de Unión al ADN/metabolismo , Glioblastoma/metabolismo , Humanos , Inmunoprecipitación/métodos , Espectrometría de Masas/métodos , Complejo Desacetilasa y Remodelación del Nucleosoma Mi-2/genética , Complejo Desacetilasa y Remodelación del Nucleosoma Mi-2/fisiología , Proteínas Nucleares/genética , Nucleosomas/genética , Unión Proteica/genética , Proteínas Represoras/fisiología , Factores de Transcripción/metabolismo , Dedos de Zinc/fisiologíaRESUMEN
Studies on activity rhythms are pivotal for the management of invasive alien species, as they provide basic insights into species basic ecology and may increase the success of control programs. The coypu Myocastor coypus, introduced from South America for fur farms, has become one of the most invasive rodents in Europe. Introduced coypus may affect crop productions, as well as natural vegetation and the breeding success of wading birds. In this study, we examined activity data collected through intensive camera-trapping in three Italian areas, including two natural areas in Northern and Central Italy, and a suburban area in Central Italy. Coypus were mostly diurnal in areas characterised by low predator pressure and, at night, they are mostly active in bright moonlight. Conversely, where predators, human pressure or numerical control programmes are present, coypus remarkably shift their behaviour towards crepuscular and night hours. In these last areas, nocturnal activity increased as moonlight decreased, possibly to reduce predation risk or encounters with humans. Where winter temperature are low, diurnal habits may have developed as a physiological adaptation and a strategy to preserve energy, potentially achieving a cost/effective thermal balance.
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The increase in size of human populations in urban and agricultural areas has resulted in considerable habitat conversion globally. Such anthropogenic areas have specific environmental characteristics, which influence the physiology, life history, and population dynamics of plants and animals. For example, the date of bud burst is advanced in urban compared to nearby natural areas. In some birds, breeding success is determined by synchrony between timing of breeding and peak food abundance. Pertinently, caterpillars are an important food source for the nestlings of many bird species, and their abundance is influenced by environmental factors such as temperature and date of bud burst. Higher temperatures and advanced date of bud burst in urban areas could advance peak caterpillar abundance and thus affect breeding phenology of birds. In order to test whether laying date advance and clutch sizes decrease with the intensity of urbanization, we analyzed the timing of breeding and clutch size in relation to intensity of urbanization as a measure of human impact in 199 nest box plots across Europe, North Africa, and the Middle East (i.e., the Western Palearctic) for four species of hole-nesters: blue tits (Cyanistes caeruleus), great tits (Parus major), collared flycatchers (Ficedula albicollis), and pied flycatchers (Ficedula hypoleuca). Meanwhile, we estimated the intensity of urbanization as the density of buildings surrounding study plots measured on orthophotographs. For the four study species, the intensity of urbanization was not correlated with laying date. Clutch size in blue and great tits does not seem affected by the intensity of urbanization, while in collared and pied flycatchers it decreased with increasing intensity of urbanization. This is the first large-scale study showing a species-specific major correlation between intensity of urbanization and the ecology of breeding. The underlying mechanisms for the relationships between life history and urbanization remain to be determined. We propose that effects of food abundance or quality, temperature, noise, pollution, or disturbance by humans may on their own or in combination affect laying date and/or clutch size.
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Nests are structures built to support and protect eggs and/or offspring from predators, parasites, and adverse weather conditions. Nests are mainly constructed prior to egg laying, meaning that parent birds must make decisions about nest site choice and nest building behavior before the start of egg-laying. Parent birds should be selected to choose nest sites and to build optimally sized nests, yet our current understanding of clutch size-nest size relationships is limited to small-scale studies performed over short time periods. Here, we quantified the relationship between clutch size and nest size, using an exhaustive database of 116 slope estimates based on 17,472 nests of 21 species of hole and non-hole-nesting birds. There was a significant, positive relationship between clutch size and the base area of the nest box or the nest, and this relationship did not differ significantly between open nesting and hole-nesting species. The slope of the relationship showed significant intraspecific and interspecific heterogeneity among four species of secondary hole-nesting species, but also among all 116 slope estimates. The estimated relationship between clutch size and nest box base area in study sites with more than a single size of nest box was not significantly different from the relationship using studies with only a single size of nest box. The slope of the relationship between clutch size and nest base area in different species of birds was significantly negatively related to minimum base area, and less so to maximum base area in a given study. These findings are consistent with the hypothesis that bird species have a general reaction norm reflecting the relationship between nest size and clutch size. Further, they suggest that scientists may influence the clutch size decisions of hole-nesting birds through the provisioning of nest boxes of varying sizes.
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This study aimed to investigate psychosocial functioning and different dimensions of theory of mind (ToM) in people with bulimia nervosa (BN) and Eating Disorder Not Otherwise Specified-BN type (EDNOS-BN). Psychosocial functioning and ToM were assessed in a sample of young adult females, 16 BN and 16 EDNOS-BN outpatients and 16 healthy controls (HCs). They were assessed using the Eating Disorder Inventory-Symptom Checklist-2 (EDI-2 SC) for evaluating psychological traits associated with eating disorders; the Symptom Checklist-90-Revised (SCL-90-R) for evaluating psychopathological status; and the Theory of Mind Assessment Scale (Th.o.m.a.s.), a semi-structured interview aimed at assessing a person's different dimensions of ToM. The BN and EDNOS-BN groups exhibited worse performance than the control group on all dimensions of the SCL-90-R, and on all dimensions of the EDI-2 SC. The only difference for perfectionism was that BN obtained higher scores than EDNOS-BN group. Our results also revealed an impairment of third-person and second-order ToM in patients with bulimia (BN and EDNOS-BN) with respect to control subjects. These preliminary data have important implications for future empirical work, in that they provide valuable information regarding the importance of investigating the various facets of ToM ability separately, in order to provide a more detailed profile of ToM functioning in the clinical samples.