bioRxiv @biorxivpreprint@qoto.org

Various bacteria are suggested to contribute to colorectal cancer (CRC) development, including pks+ E. coli which produce the genotoxin colibactin that induces characteristic mutational signatures in host epithelial cells. It remains unclear how the highly unstable colibactin molecule is able to access host epithelial cells and its DNA to cause harm. Using the microbiota-dependent ZEB2-transgenic mouse model of invasive CRC, we found that pks+ E. coli drives CRC exacerbation and tissue invasion in a colibactin-dependent manner. Using isogenic mutant strains, we further demonstrate that CRC exacerbation critically depends on expression of the E. coli type-1 pilus adhesin FimH and the F9-pilus adhesin FmlH. Blocking bacterial adhesion using a pharmacological FimH inhibitor attenuates colibactin-mediated genotoxicity and CRC exacerbation. Together, we show that the oncogenic potential of pks+ E. coli critically depends on bacterial adhesion to host epithelial cells and is critically mediated by specific bacterial adhesins. Adhesin-mediated epithelial binding subsequently allows production of the genotoxin colibactin in close proximity to host epithelial cells, which promotes DNA damage and drives CRC development. These findings present promising therapeutic avenues for the development of anti-adhesive therapies aiming at mitigating colibactin-induced DNA damage and inhibiting the initiation and progression of CRC, particularly in individuals at risk for developing CRC. ### Competing Interest Statement The authors have declared no competing interest.

Prostate adenocarcinoma (PRAD) is the second most frequent cancer among men worldwide. lncRNAs have been suggested as novel promising biomarkers for cancer diagnosis and prognosis and therapeutic targets. Despite the previous investigations, still, comprehensive studies are required to discover the regulatory roles of lncRNAs in PRAD. We introduced a PRAD lncRNA-mRNA network (PLMNET) to identify novel PRAD biomarkers and therapeutic targets among lncRNAs. The PLMNET is constructed based on RNAs that are differentially expressed and had strongly correlate with each other. PANTR1, HOXB-AS3, EMX2OS, GATA3-AS1, LINC01116, LINC02385 are PLMNET hubs that are considered key regulatory lncRNAs of PRAD. We also mapped PLMNET lncRNAs to their associated GO-terms and hallmarks. Literature review confirms our suggested biomarkers and their associated hallmarks. The high degree nodes of PLMNET, GATA3-AS1, EMX2OS, LINC01116, LINC02137, RP11-431N15.2, LINC01671, LINC02806, ENSG00000231196, ENSG00000244252, RP11-379F12.3, and SLC2A9-AS1 indicate significant association with overall survival. In addition, we found that the hubs regulatory roles have previously been confirmed in different types of cancers by literature. For example, PANTER hub exhibited a high expression correlation with three known tumor suppressors (SKI, SKP2, and SH2B3) and is associated with Sustaining proliferative signal. The literature has confirmed that PANTER may promote cancer cell proliferation in prostate carcinoma. It also promotes renal cell carcinoma and Hepatocellular carcinoma progression. Furthermore, literature review proves that EMX2OS and LINC01116 are key regulatory lncRNAs of prostate adenocarcinoma and HOXB-AS3 and GATA3-AS1 have regulatory roles in other cancer. According to pathway analysis, PLMNET hubs seem to contribute to PRAD proliferation, invasion, metastasis, and apoptosis via dysregulation of anaphase promoting complex/cyclosome (APC/C) subunits and dysregulation of mitotic cell cycle. These results introduce PLMCNET hubs as potential molecular biomarkers for diagnosis and prognosis of PRAD and suggest therapeutic targets for PRAD patients. ### Competing Interest Statement The authors have declared no competing interest.

Background: As biological data increases, we need additional infrastructure to share it and promote interoperability. While major effort has been put into sharing data, relatively less emphasis is placed on sharing metadata. Yet, sharing metadata is also important, and in some ways has a wider scope than sharing data itself. Results: Here, we present PEPhub, a new tool focused on improving sharing and interoperability of biological metadata. PEPhub provides an API, natural language search, and user-friendly web-based sharing and editing of sample metadata tables. These features will help researchers to share biological metadata more broadly. Availability: https://pephub.databio.org ### Competing Interest Statement NCS is a consultant for InVitro Cell Research, LLC.

Partial least squares regression (PLSR) is a reference method in chemometrics. In agronomy, it is used for instance to predict components of chemical composition (response variables y) of vegetal materials from spectral near infrared (NIR) data X collected from spectrometers. The principle of PLSR is to reduce the dimension of the spectral data X by computing vectors that are then used as latent variables (LVs) in a multiple linear model. A difficulty is to determine the relevant dimensionality (number of LVs) of the model for the given available data. This step can also become time consuming when many different datasets have to be processed and/or the datasets are frequently updated. An alternative to determinate the relevant PLSR dimensionality is the ensemble learning method 'PLSR-averaging'. In the past, this method has been demonstrated to be efficient for complex biological materials such as mixed forages, and facilitates to automatize predictions (e.g. in user-friendly web interface platforms). This article presents the extension of the PLSR-averaging to a k-nearest neighbors locally weighted PLSR pipeline (kNN-LWPLSR). The kNN-LWPLSR pipeline has the advantage to account for non-linearity between X and y existing for instance in heterogeneous data (e.g. mixing of vegetal species, collection from different geographical areas, etc.). In the article, kNN-LWPLSR-averaging is applied to an extensive NIR database built to predict the chemical composition of European and tropical forages and feed. The main finding of the study was the overall superiority of the averaging compared to the usual kNN-LWPLSR. Averaging may therefore be recommended in local-PLSR pipelines to predict NIR forage data. ### Competing Interest Statement The authors have declared no competing interest.

Drought stress poses a significant threat to crop productivity, making the development of drought-tolerant crops a priority. The impact of drought on grain yield loss varies significantly, ranging from 10% to 76%, depending on the specific stage of occurrence and the severity of the drought. In this study, we investigated the effects of introducing the pSARK::IPT transgene on the drought tolerance and nutritional composition of successive generations of tropical maize. Towards this goal, we screened different generations of maize plants by genotyping PCR, exposed them to long-term drought stress and analysed several drought stress markers and nutritional profiles of the plants. Our results demonstrated that the pSARK::IPT transgene was present in 4 successive generations of maize plants. Under drought conditions, transgenic maize exhibited higher relative water content, and delayed senescence compared to wild-type plants. Additionally, transgenic plants showed increased levels of total chlorophyll, chlorophyll a, and chlorophyll b, indicating improved photosynthetic activity under water deficit. Our study also showed that IPT-transgenic plants produced substantially higher yields and demonstrated enhanced nutritional value compared to wild-type plants when grown under well-watered conditions. Further research is warranted to investigate the underlying molecular mechanisms involved in these improvements and assess the performance of pSARK::IPT maize under field conditions. ### Competing Interest Statement The authors have declared no competing interest.

Humans are a major evolutionary force on wildlife via artificial selection. While often explored through the lens of extractive interactions (e.g., hunting) able to favour certain behavioural traits over others, the implications of non-extractive ones, such as wildlife feeding, remain under-studied. Research has recently shown that people tend to feed (and sometimes favour) a limited subset of bolder individuals within natural populations, although its dynamics and consequences are not fully clear. Using fallow deer living in a peri-urban setting as a model population, we studied whether mother deer that display reduced fear of humans and consistently approach them for food adopt weaker anti-predator strategies by selecting for fawning bedsites that are less concealed and closer to human hotspots, allowing them to take advantage of additional artificial feeding opportunities in comparison to shier mothers in this population. Our dataset encompassed 171 fawns from 109 mothers across 4 years. Contrary to our expectations, we found that mothers that regularly accepted food from humans selected for more concealed bedsites farther away from them, giving their offspring better protection while also taking advantage of additional artificial food during lactating. Our results show marked behavioural adaptation by a subset of females, making this the first time that the link between tendency to approach humans and strategies to protect offspring is explored. Given previous findings that these begging females also deliver heavier fawns at birth, our research adds a piece to the complex puzzle describing human manipulation of behaviour in natural populations and its fitness consequences. ### Competing Interest Statement The authors have declared no competing interest.

Arctic coastal ecosystems include benthic communities that hold an important role within the marine food chain. Kelps, fucoid species, and coralline algae dominate rocky habitats, offering food and shelter for various species. Kelps and fucoid species also aid in carbon sequestration, sediment stabilization, and erosion mitigation. In summer, the influx of freshwater from glacier and permafrost melt alters coastal waters conditions. The input of turbid freshwater influences underwater light, salinity, and substrate, impacting benthic organism distribution. This study investigates possible link between environmental conditions and benthic diversity through environmental DNA (eDNA). Six sites were monitored along Kongsfjorden (Svalbard, Norway) during the summer of 2021. Contrary to expectations, macroalgal distribution didn't correlate with light, and suspension feeders showed no clear links with chlorophyll a or nutrient concentrations. Glacial influence may have contributed to higher benthic diversity. Predators' presence, tied to glacier proximity, possibly explained this trend. However, further studies are needed to validate these observations and assumptions. ### Competing Interest Statement The authors have declared no competing interest.

Biodiversity loss presents a growing threat to the global environment and requires systematic and spatially contiguous monitoring. Monitoring of within-species genetic variation, a key factor when assessing biodiversity loss, is laborious and could be complemented by remote observations of phenotypes allowing inferences about genetic variation. We studied genetic and phenotypic variations in the common European beech (Fagus sylvatica L.) derived from whole-genome sequences and spectral phenotypes of more than 200 individuals at 22 sites across the species' natural range. The spectral phenotypes were collected under standardized illumination/observation conditions from the same top-of-canopy leaves used for nuclear DNA extraction. We found that information from spectral phenotypes positively contributes to predictions of genetic structure using environmental variables across the sites in our dataset. We therefore propose that future spaceborne satellites with sufficient spectral and spatial resolution, repeatedly acquiring spectral phenotypes globally across species ranges, improve our ability to rapid and spatially contiguous monitoring of genetic diversity in trees. ### Competing Interest Statement The authors have declared no competing interest.

The phytochrome (phy) family of sensory photoreceptors modulates developmental programs in response to ambient light. phys control gene expression in part by directly interacting with the bHLH class of transcription factors, phytochrome-interacting factors (PIFs), and inducing their rapid phosphorylation and degradation. Several kinases have been shown to phosphorylate PIFs and promote their degradation. However, the phosphatases that dephosphorylate PIFs are less investigated. Here we describe the identification of four regulatory subunits of Arabidopsis protein phosphatase 2A (PP2A) family (B'α, B'β, B''α and B''β) that interacted with PIF3 in yeast-two-hybrid, in vitro and in vivo assays. The pp2ab''αβ and b''αβ/b'αβ displayed short hypocotyls, while the overexpression of the B subunits induced longer hypocotyls compared to wild type under red light. The light-induced degradation of PIF3 was faster in b''αβ/b'αβ quadruple mutant compared to wild type. Consistently, immunoprecipitated PP2A A and B subunits directly dephosphorylated PIF3-MYC in vitro. RNA-seq analyses showed that B''α and B''β alter global gene expression in response to red light. PIFs (PIF1, PIF3, PIF4 and PIF5) are epistatic to these four B subunits in regulating hypocotyl elongation under red light. Collectively, these data show an essential function of PP2A in dephosphorylating PIF3 to modulate photomorphogenesis in Arabidopsis.

The anadromous Atlantic salmon undergo a preparatory physiological transformation before seawater entry, referred to as smoltification. Key molecular developmental processes involved in this life stage transition, such as remodeling of gill functions, are known to be synchronized and modulated by environmental cues like photoperiod. However, little is known about the photoperiod influence and genome regulatory processes driving other canonical aspects of smoltification such as the large-scale changes in lipid metabolism and energy homeostasis in the developing smolt liver. Here we generate transcriptome, DNA methylation, and chromatin accessibility data from salmon livers across smoltification under different photoperiod regimes. We find a systematic reduction of expression levels of genes with a metabolic function, such as lipid metabolism, and increased expression of energy related genes such as oxidative phosphorylation, during smolt development in freshwater. However, in contrast to similar studies of the gill, smolt liver gene expression prior to seawater transfer was not impacted by photoperiodic history. Integrated analyses of gene expression and transcription factor (TF) binding signatures highlight likely important TF dynamics underlying smolt gene regulatory changes. We infer that ZNF682, KLFs, and NFY TFs are important in driving a liver metabolic shift from synthesis to break down of organic compounds in freshwater. Moreover, the increased expression of ribosomal associated genes after smolts were transferred to seawater was associated with increased occupancy of NFIX and JUN/FOS TFs proximal to transcription start sites, which could be the molecular consequence of rising levels of circulating growth hormones after seawater transition. We also identified differential methylation patterns across the genome, but associated genes were not functionally enriched or correlated to observed gene expression changes across smolt development. This contrasts with changes in TF binding which were highly correlated to gene expression, underscoring the relative importance of chromatin accessibility and transcription factor regulation in smoltification. ### Competing Interest Statement The authors have declared no competing interest.

Conifers are long-lived and slow-evolving, thus requiring effective defences against their fast-evolving insect natural enemies. The copy number variation (CNV) of two key acetophenone biosynthesis genes Ugt5/Ugt5b and Betaglu-1 may provide a plausible mechanism underlying the constitutively variable defence in white spruce (Picea glauca) against its primary defoliator, spruce budworm. This study develops a long-insert sequence capture probe set (Picea\_hung\_p1.0) for quantifying copy number of Betaglu-1-like, Ugt5-like genes and single-copy genes on 38 Norway spruce (Picea abies) and 40 P. glauca individuals from eight and nine provenances across Europe and North America respectively. We developed local assemblies (Piabi\_c1.0 and Pigla\_c.1.0), full-length transcriptomes (PIAB\_v1 and PIGL\_v1), and gene models to characterise the diversity of Betaglu-1 and Ugt5b genes. We observed very large copy numbers of Betaglu-1, with up to 381 copies in a single P. glauca individual. We observed among-provenance CNV of Betaglu-1 in P. glauca but not P. abies. Ugt5b was predominantly single-copy in both species. This study generates critical hypotheses for testing the emergence and mechanism of extreme CNV, the dosage effect on phenotype, and the varying copy number of genes with the same pathway. We demonstrate new approaches to overcome experimental challenges in genomic research in conifer defences. ### Competing Interest Statement The authors have declared no competing interest.

Urbanization alters many biotic and abiotic environmental factors, but the effects of urbanization on local adaptation and coevolution between species remains largely unstudied. Using a common garden experiment of 30 populations and 1080 plants, we tested for local adaptation in the mutualism between white clover (Trifolium repens) and rhizobia (Rhizobium leguminosarum symbiovar trifolii) along an urbanization gradient. Our results show a spatial mosaic in local adaptation, with stronger local adaptation for rhizobia than white clover. While nitrogen addition (N) strongly influenced plant biomass and nodule density, it did not consistently mediate patterns of local adaptation. The strength of local adaptation was positively related between white clover and rhizobia under N addition, suggesting that soil N mediates coadaptation between white clover and rhizobia. Local adaptation was not influenced by urbanization, indicating that while urbanization does influence the ecology of plant-microbe interactions, it does not disrupt local adaptation and coevolution among mutualists. ### Competing Interest Statement The authors have declared no competing interest.

Large lipid-storing copepods dominate mesozooplankton biomass in the polar oceans and form a critical link between primary production and higher trophic levels. The ecological success of these species depends on their ability to survive periods of food deprivation in a highly seasonal environment, but the molecular changes that mediate starvation tolerance in these taxa are unknown. We conducted starvation experiments for two dominant Southern Ocean copepods, Calanoides acutus and Calanus propinquus, allowing us to compare the molecular starvation response between species. These species differ in life history, diet, and metabolic traits, and expressed overlapping but distinct transcriptomic responses to starvation. Most starvation-response genes were species-specific, but we identified a conserved core set of starvation-response genes related to RNA and protein metabolism. We used phylotranscriptomics to place these results in the context of copepod evolution and found that starvation-response genes are under strong purifying selection at the sequence level and stabilizing selection at the expression level, consistent with their role in mediating essential biological functions. Selection on starvation-response genes was especially strong in our focal lipid-storing lineage relative to other copepod taxa, underscoring the significance of starvation tolerance for these species. We also found that certain key lipid enzymes (elongases and desaturases) have experienced diversification and positive selection in lipid-storing lineages, reflecting the unique lipid storage needs of these animals. Our results shed light on the molecular adaptations of high-latitude zooplankton to variable food conditions, and suggest that starvation-response genes are under particularly strong sequence and expression constraints. ### Competing Interest Statement The authors have declared no competing interest.

The emergence and subsequent diversification of morphological novelties is a major feature of animal evolution1–9. However, in most cases little is known about the molecular basis of the evolution of novel structures and the genetic mechanisms underlying their diversification. The epandrial posterior lobes of the male genital arch is a novelty of some species of the Drosophila melanogaster subgroup10–13. The posterior lobes grasp the ovipositor of the female and then integrate between her abdominal tergites, and therefore these structures are important for copulation and species-recognition10–12,14–17. The posterior lobes evolved from co-option of a Hox regulated gene network from the posterior spiracles10 and have since diversified in shape and size in the D. simulans clade in particular over the last 240,000 years driven by sexual selection18–21. The genetic basis of this diversification is highly polygenic but to the best of our knowledge none of the causative genes have yet been identified despite extensive mapping22–30. Identifying the genes underlying the diversification of these secondary sexual structures is essential to understanding the basis of changes in their morphology and the evolutionary impact on copulation and species recognition. Here, we show that the transcription factor encoded by Sox21b negatively regulates posterior lobe size during development. This is consistent with higher and expanded expression of Sox21b in D. mauritiana, which develops smaller posterior lobes compared to D. simulans. We tested this by generating reciprocal hemizygotes and confirmed that changes in Sox21b underlie posterior lobe evolution between these two species. Furthermore, we found that differences in posterior lobe size caused by the species-specific allele of Sox21b significantly affect the duration of copulation. Taken together, our study reveals the genetic basis for the sexual selection driven diversification of a novel morphological structure and its functional impact on copulatory behaviour. ### Competing Interest Statement The authors have declared no competing interest.

Challenging or adverse early-life conditions, even when transient, can have long-lasting effects on individual phenotypes and reduce lifespan across species. If these effects can be mitigated, even in part, by a high quality later-life environment, then differences in future resource access may explain variation in vulnerability and resilience to early-life adversity. Using 32 years of data on 886 wild North American red squirrels, we test the hypothesis that the negative effects of early-life adversity on lifespan can be mitigated by later-life food abundance. We first define early-life adversities as factors that significantly reduce the likelihood of juvenile survival, and find that they had cumulative negative effects on lifespan. We then show that although experimental supplementation with additional food increases individual lifespan, it did not change the consequences of early-life adversity on longevity. A naturally-occurring future food boom experienced in the second year of life, however, did eliminate the longevity costs of a harsh early-life environment. Together, our results demonstrate that adverse conditions experienced early in life reduce lifespan in red squirrels and thus may influence patterns of natural selection beyond juvenile viability. That these effects can be mitigated by a high-quality future environment suggests a non-deterministic role for early-life conditions on later-life phenotypes, and highlights the importance of evaluating the impact of early-life conditions in the context of an animals entire life course. ### Competing Interest Statement The authors have declared no competing interest.

Grapevine downy mildew, caused by the oomycete Plasmopara viticola, is a globally destructive disease that particularly affect the Eurasian wine grape Vitis vinifera. While genetically resistant varieties are becoming more accessible, populations of P. viticola are demonstrating rapid adaptability, successfully overcoming these resistances. Here we aimed to identify the avirulence genes involved in the interaction with the Rpv3.1-mediated resistance in grapevine. We sequenced the full genome of 136 P. viticola strains sampled in a natural population of Bordeaux (France) and characterized their development on both resistant and sensitive cultivars. The genome-wide association study allowed the identification of a genomic region associated with the breakdown of Rpv3.1 grapevine resistance (avrRpv3.1 locus). A diploid-aware reassembly of the P. viticola INRA-Pv221 genome allowed to detect structural variations in this locus, including a major 30 Kbp deletion. At the avrRpv3.1 locus, virulent P. viticola strains presented deletion on both haplotypes indicating that avirulence is recessive. The deletion involves two closely-related genes that encode proteins containing 800-900 amino acids with a signal peptide. The structure of the predicted proteins contains repeats of the LWY-fold structural modules, typical of oomycete effectors. Moreover, when these proteins were transiently expressed, they induced cell death in grapevines carrying Rpv3.1 resistance, confirming their avirulence nature. The first description of candidate effectors of P. viticola involved in the interaction with resistance gene provides valuable insights into the genetic mechanisms that enable P. viticola to adapt to grapevine resistance, laying a foundation for developing strategies to manage this damaging crop pathogen. ### Competing Interest Statement The authors have declared no competing interest.

Insecticide resistance is a serious threat to our ability to control mosquito vectors which transmit pathogens including malaria parasites and arboviruses. Understanding the underlying mechanisms is an essential first step in tackling the challenges presented by resistance. This study aimed to functionally characterise the carboxylesterase, CCEae3A, the elevated expression of which has been implicated in temephos resistance in Aedes aegypti and Aedes albopictus larvae. Using our GAL4/UAS expression system, already established in insecticide-sensitive Anopheles gambiae mosquitoes, we produced transgenic An. gambiae mosquitoes that express an Ae. aegypti CCEae3A ubiquitously. This new transgenic line permits examination of CCEae3A expression in a background which does not express the gene and allows comparison with existing An. gambiae GAL4-UAS lines. Insecticide resistance profiling of these transgenic An. gambiae larvae indicated significant increases in resistance ratio for three organophosphate insecticides, temephos (5.98), chloropyriphos (6.64) and fenthion (3.18) when compared to the parental strain. Cross resistance to adulticides from three major insecticide classes: organophosphates (malathion, fenitrothion and pirimiphos methyl), carbamates (bendiocarb and propoxur) and pyrethroid (alpha-cypermethrin) was also detected. Resistance to certain organophosphates and carbamates validates conclusions drawn from previous expression and phenotypic data. However, detection of resistance to pirimiphos methyl and alphacypermethrin has not previously been formally associated with CCEae3A, despite occurring in Ae. aegypti strains where this gene was upregulated. Our findings highlight the importance of characterising individual resistance mechanisms, thereby ensuring accurate information is used to guide future vector control strategies. ### Competing Interest Statement The authors have declared no competing interest.

Biobanking of tissue from clinically obtained kidney biopsies for later use with multi-omic and imaging techniques is an inevitable step to overcome the need of disease model systems and towards translational medicine. Hence, collection protocols ensuring integration into daily clinical routines using preservation media not requiring liquid nitrogen but instantly preserving kidney tissue for clinical and scientific analyses of paramount importance. Thus, we modified a robust single nucleus dissociation protocol for kidney tissue stored snap frozen or in the preservation media RNAlater and CellCover. Using porcine kidney tissue as surrogate for human kidney tissue, we conducted single nucleus RNA sequencing with the Chromium 10X Genomics platform. The resulting data sets from each storage condition were analyzed to identify any potential variations in transcriptomic profiles. Furthermore, we assessed the suitability of the preservation media for additional analysis techniques (proteomics, metabolomics) and the preservation of tissue architecture for histopathological examination including immunofluorescence staining. In this study, we show that in daily clinical routines the RNAlater facilitates the collection of highly preserved kidney biopsies and enables further analysis with cutting-edge techniques like single nucleus RNA sequencing, proteomics, and histopathological evaluation. Only metabolome analysis is currently restricted to snap frozen tissue. This work will contribute to build tissue biobanks with well-defined cohorts of the respective kidney disease that can be deeply molecularly characterized, opening new horizons for the identification of unique cells, pathways and biomarkers for the prevention, early identification, and targeted therapy of kidney diseases. ### Competing Interest Statement The authors have declared no competing interest.

Evolutionary transitions in individuality (ETIs), such as the emergence of multicellularity, are events in the history of life during which entities at one level of organisation (particles) form collective-level entities that subsequently become individuals in their own right. Recent empirical and theoretical studies advocate the importance of an externally imposed meta-population structure or "ecological scaffold" for the emergence of new levels of individuality. Such a scaffold enables survival and reproduction at the collective level and thus the possibility of selection for beneficial traits on that level. However, a longstanding difficulty for the ecological scaffolding approach has been its inability to adequately explain how collective-level trait values that evolved under scaffolding conditions can be retained once these conditions are lifted. We call this difficulty the problem of endogenisation. Here, we derive general conditions for the possibility of endogenisation. Key to endogenisation is the existence of a fitness valley that can be circumvented when scaffolding occurs. Using a stochastic meta-population model, we implement two versions of ecological scaffolding (one temporal and one spatial) and study subsequent evolutionary trajectories using the modelling techniques of adaptive dynamics. Our analysis yields several important results. The temporal model reveals that only collective traits based on particle-particle interactions can be endogenised when a temporary scaffold is applied to the entire population. The spatial model shows that, given the presence of an environmental gradient of externally imposed meta-population structure, ecological scaffolding can only occur in a limited "Goldilocks" zone of the environment. Further, if endogenisation conditions are also fulfilled, scaffolded collectives can colonise non-scaffolding areas of the environment. We conjecture that Goldilocks zones could act as initiators of ETIs and help explain the near ubiquity of collective-level individuality even if the conditions that promote it prove to be rare. ### Competing Interest Statement The authors have declared no competing interest.

The dopamine transporter (DAT) plays a crucial role in regulating the brain's dopamine (DA) homeostasis. Atypical DAT deficiency syndrome (DTSD) is a disease characterized by early-onset parkinsonism and comorbid psychiatric symptoms, but the pathobiological processes that link DAT dysfunction to both parkinsonism and psychiatric symptoms are unknown. Here, we present a genetic mouse model of atypical DTDS that expresses two coding DAT variants, DAT-I312F and DAT-D421N, derived from a patient diagnosed with ADHD and parkinsonism. Phenotypic characterization of the mutant mice revealed impaired DAT function and major homeostatic changes including increased ambient extracellular DA levels, decreased evoked DA release, and reduced expression of both tyrosine hydroxylase (TH) and of DA D1/D2 receptors. This was accompanied by diminished striatal dopaminergic axonal density and a psychomotor phenotype characterized by hyperactivity, enhanced exploratory activity, and pronounced clasping. Importantly, both amphetamine and anticholinergic treatment ameliorated aberrant hyperlocomotion in the mice. Summarized, by replicating core aspects of the patient's phenotype, the mouse model not only provides insights into the mechanisms underlying atypical DTDS but also underlines the broad relevance of DA deficits for understanding the co-morbidity between neuropsychiatric diseases and parkinsonism ### Competing Interest Statement The authors have declared no competing interest.