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Diversity at the HYP1 locus in potato cyst nematodes does not result from developmentally-programmed somatic mutations.

Most genetic diversity stems from spontaneous mutations, that is, errors in DNA repair or replication. But for dozens of organisms across the tree of life, mutations at specific loci are not spontaneous but developmentally programmed: effectively, some organisms edit their own DNA sequences. This is perhaps most common among pathogens and parasites, many of which use editing to diversify genes that produce important antigens. Plant-parasitic potato cyst nematodes are damaging agricultural pests that establish a lifelong feeding site inside the root of their host plant. We previously observed extensive diversity of rare alleles at HYP1, the most highly expressed gene that encodes a protein secreted by potato cyst nematodes during parasitism. Importantly, HYP1 alleles differ from each other by complex, in-frame rearrangements of short repeated sequence motifs within a single exon. Combining several lines of evidence, we previously hypothesized that potato cyst nematodes use developmentally-programmed mutations, or editing, to diversify HYP1 alleles in the soma. In the current work, we now test this hypothesis. We employ highly accurate long-read DNA sequencing of a simplified genetic system to identify potential rare edited alleles, we use a transgenic yeast system to describe large de novo mutations at HYP1, and we interpret our findings in light of key population genetic parameters as well as the genetic diversity surrounding HYP1 and across the genome.

Animals

The maternal-to-zygotic transition is a critical window for PFOA-induced disruption of developmental programming.

Early embryogenesis is governed by precisely timed gene regulatory programs that coordinate cell fate specification, tissue patterning, and morphogenesis. The maternal-to-zygotic transition (MZT) represents a pivotal developmental milestone during which regulatory control shifts from maternally deposited transcripts to activation of the zygotic genome. Disruption of this transition has the potential to alter developmental trajectories with lasting consequences. Per- and polyfluoroalkyl substances (PFAS), environmentally persistent contaminants, have been linked to developmental abnormalities, yet their impact on core embryonic gene regulatory networks especially with exposure during MZT is not well understood. Using zebrafish (Danio rerio), a tractable vertebrate model and New Approach Methodology (NAM), we investigated how PFAS exposure during the MZT alters early developmental programming. Embryos were exposed starting at different times before and within the MZT time window and collected at 24 h post-fertilization (hpf) for transcriptomic analysis. Targeted qRT-PCR revealed dysregulation of genes controlling transcriptional activation, lineage specification, proliferation, and differentiation. Whole-transcriptome RNA sequencing (RNA-seq) further identified widespread perturbations in gene networks governing transcriptional regulation, cell signaling, and embryonic morphogenesis. Temporal analysis revealed that exposure beginning at 3.5 hpf, followed by 8 hpf, corresponding to early zygotic genome activation and near completion of zygotic activation, respectively, resulted in the greatest differential gene expression changes at 24 hpf. Consistent with these early gene regulatory perturbations, larvae exposed starting at 8 hpf also exhibited altered behavior at 5 days post-fertilization. Together, these findings demonstrate that PFAS exposure during MZT disrupts the establishment of embryonic gene regulatory networks, linking environmental toxicant exposure to altered developmental patterning and organismal outcomes. This work underscores the vulnerability of early developmental transitions to environmental perturbation and positions MZT as a critical window of susceptibility during development.

NAMs (new approach methodologies)

[Selection and characterization of Penicillium cyclopium mutants with altered developmental program].

After treatment with NNMG, NaNO2 or UV-light of conidiospores of Penicillium cyclopium strain SM 72 variant strains (mutants) with altered developmental programme were selected. Additionally from a methionine-auxotrophic mutant of P. cyclopium prototrophic revertants were prepared. Investigation of the alkaloid metabolism and other idiophase processes has shown that these mutant strains can be divided into two groups (cf. table 1): a) Mutants with a depression of all idiophase features. The defects of these strains presumably affect central regulatory processes which, as in strain rev-met 83a, can be reversed spontaneously by an one-step mechanism and b) mutants defective in certain parts of the idiophase programme only, demonstrating that there is a certain autonomy in regulation of the individual parts of the programme.

Alkaloids

Genetic determination of the alpha-galactosidase developmental program in mice.

The expression of alpha-galactosidase in liver, heart, and brain during postembryonic development has been examined in several inbred mouse strains. In most strains, the developmental patterns of alpha-galactosidase are coordinate with those of two other acid hydrolases, beta-glucuronidase and beta-galactosidase. Certain inbred mouse strains, including members of the C57-C58 family, have a tissue-specific alteration in the temporal expression of alpha-galactosidase activity. This alteration shows additive inheritance and appears to be controlled by a single genetic locus. The altered developmental expression of the enzyme is not accompanied by any discernible change in its physical properties.

Animals

Developmental restrictions on transcription: determinants of the developmental program and their role in aging.

A developing plant system, the soybean hypocotyl has been used to investigate early transcription events which restrict auxin induced cellular proliferation to the appropriate developmental stage. Auxin treatment of 4-day old seedlings resulted in an early (6 hour) activation of chromatin-bound RNA polymerase activity wihich approached 200% of control values by 18 hours. This occurred without a detectable alteration in chromatin template capacity (assayed with exogenous RNA polymerase) and resulted in the synthesis of "induced RNA transcripts" as determined in vitro by nearest neighbor analysis. In contrast, auxin treatment of unresponsive 8-day old seedlings did not alter the chromatin-bound RNA polymerase activity. Hormonal activation did, however, result in the exposure of "induced template" regions in chromatin which could only be transcribed in vitro if exogenous RNA polymerase was included in the transcription reaction. Isoelectric focusing of the endogenous chromatin-bound and soluble RNA polymerase enzymes from successive developmental stages revealed that the chromatin-bound enzymes at the 2-day stage were first released from the chromatin complex and could be recovered in the soluble pool (4-day stage). This was followed by a gradual disappearance of these subspecies (6-day stage) until only a limited ensemble of RNA polymerase subspecies remained bound to chromatin and free in the soluble pool (8-day stage). Similar analyses of both the bound and free enzymes at the 4 and 8-day stages following auxin treatment revealed that the 4-day soluble enzymes could be induced to rebind to the chromatin complex in a defined sequence after hormone treatment while those of the 8-day hypocotyl were unable to do so. These developmental events indicate that the select loss of certain RNA polymerase subspecies serves to restrict the hormone responsivness of this tissue to the early developmental stages. Such restrictions could thus commit the constituent hypocotyl cells to their terminal post-mitotic phase of development.

2,4-Dichlorophenoxyacetic Acid

Genetic determination of the developmental program for mouse liver beta-galactosidase: involvement of sites proximate to and distant from the structural gene.

The identification and mode of action of genetic loci that program gene expression during development are important for understanding differentiation in higher organisms. Previous work from this laboratory has identified two patterns for the postnatal development of liver beta-galactosidase among inbred mouse strains: type I, where activity levels remain constant after about 30 days of age, is found in strains DBA/2J, CBA/J, and BALB/cJ, among others; type II, where activity levels increase between 25 and 50 days of age to reach a new adult level, is found in strain C57BL/6J and related strains. It has been shown that the type I vs. type II developmental difference between strains C57BL/6J and DBA/2J is due to variation at a locus, Bgl-t, that maps with the beta-galactosidase complex, [Bgl], on chromosome 9. In the present study, we have confirmed the existence of Bgl-t as a temporal locus within [Bgl] by analysis of both a congenic strain carrying the beta-galactosidase complex of strain CBA/J in the C57BL/6J genetic background and a cross of strains CBA/J and C57BL/6J. The existence of additional temporal loci for beta-galactosidase that segregate independently of the structural gene and participate in determination of the type I vs. type II difference was revealed by analysis of: (1) a congenic strain containing the beta-galactosidase complex of strain BALB/cJ in the C57BL/10Sn background; (2) recombinant inbred lines derived from progenitor strains C57BL/6ByJ and BALB/cByJ; and (3) a genetic cross between strains C57BL/6ByJ and BALB/cByJ. Thus, for these pairs of strains, the type I vs. type II developmental difference is due to variation at a temporal locus (or loci) unlinked to the enzyme structural gene, and not at Bgl-t. These facts, together with information gathered from an examination of the distribution of beta-galactosidase phenotypes among over 100 inbred strains (Breen, Lusis and Paigen 1977), have led us to conclude that the postnatal developmental pattern for liver beta-galactosidase is determined by a set of interacting temporal genes. One of these, Bgl-t, is located within [Bgl], and one or more are separable from [Bgl] by recombination. A possible mode of interaction among the temporal and instructural loci is suggested.

Animals

Developmental program of murine erythroleukemia cells. Effect of the inhibition of protein synthesis.

The relationship between protein synthesis and commitment to terminal erythroid differentiation by dimethylsulfoxide-treated murine erythroleukemia (MEL) cells has been studied. Treatment with cycloheximide blocks the commitment of MEL cells. The effects of cycloheximide are completely reversible, however. Treatment of MEL cells before commitment delays commitment for a period of time equal to the length of inhibitor treatment. Puromycin exerts a similar effect on the commitment of MEL cells. These results indicate that there is a continuous requirement for protein synthesis before the commitment event.

Animals

Developmental programming for retinotectal patterns.

Programming events at stages 28-31 in Xenopus specify the cellular positional information that individual retinal ganglion cells will use to derive theri appropriate locus specificity for assembly of the retinotectal map. The 'programme' that emerges in the stage 31 retina affects the entire ganglion cell population (99 percent of which is generated later) and refers positional information to intraretinal axes (AP and DV) and zero-points. Its expression in intact retinae was not modified by repeated reintroduction into pre-stage 28 orbits, prolonged eye culture in vitro, or severe disruption of the timing and sequence of ganglion cell births or of optic fibre arrivals in the tectum. In contrase, intraretinal reorganizations did not produce major modifications (e.g. after transection, partial ablation, fragment fusion etc.) in the set of locus specificities arising in various retinal regions and, in some instances, in the reference axes themselves. The modified programmes were characteristic of the components undergoing reorganization, but were convergent (many sets of reorganizing components gave a few final patterns); they appeared to involve a stable and rapid reprogramming of certain components by others, in a hierarchical fashion. The remaining experiments focus on the problem of localizing the 'trigger' for the transition from the unspecified to the specified state in the retina at stages 28-31. Specification can occur in vitro, based on reversible AP and DV orientational markers which are present in the pre-stage 28 eye primordium; in heterochromically grafted eyes, specification was neither precipitated nor delayed by altering the stage of the host. Finally, chemical dissection of the differentiating eye primordium confirmed the inference (from Jacobson's [3-H] thymidine labelling kinetics) that a specific gangliogenic precursor cell type exists in the stage 28 retinal neuroepithelium, and suggested that differentiative events in these cells trigger the specification process.

Animals

Interdisciplinary early intervention program.

Developmental data were obtained on 40 children with Down's syndrome by an interdisciplinary team during an ongoing early intervention program. Interventive methods are described. Results are compared with available data on children with Down's syndrome who were not in early intervention programs. Comparisons of developmental progress are also made of children begun in the early intervention program before six months of age with those begun after six months of age. Conclusions from the study indicate that early intervention helps the child in earlier attainment of many developmental tasks and enhances functioning of the family unit.

Counseling

The area-code hypothesis: the immune system provides clues to understanding the genetic and molecular basis of cell recognition during development.

Numberous studies of embryogenesis have provided evidence for highly specific cell-surface recognition phenomena. These include both the interactions of neighboring cells and the specific cellular migrations which occur as the developmental program of the embryo progresses. The area-code hypothesis elaborate here is an attempt to provide a framework for understanding cell-recognition phenomena in development. This hypothesis is based on extensive genetic, molecular, and cellular studies of the immune system. These studies suggest that the following events occur during the differentiation of antibody-producing cells. 1) Somatic cell lines of antibody-producing cells undergo a modification of their DNA as they become committed to synthesize a particular type of antibody molecule. This chromosomal modification event is probably a DNA translocation which leads to a somatic rearrangement of certain antibody genes. 2) In each of the specific cell lineages the new arrangement of DNA is inherited by all subsequent generations of cells. 3) The developmental programs which control these genetic alterations may be employed in a programmed and reproducible fashion. This programming of antibody development is suggested because different embryos appear to become committed to the production of identical antibody molecules in the same developmental sequence. 4) Antibody molecules are initially displayed on the cell surface where they serve as highly specifici receptors to trigger the cell to proliferate and differentiate upon interacting with appropriate external molecular signals. 5) Antibody-producing cells display combinations of different molecules on their surfaces which cause each of a very large number of different cells to interact differently with their environment. 6) The genes which code for many of these cell-surface molecules are organized into multigene families. These observations as well as information from other developmental systems have led us to propose the area-code hypothesis. This hypothesis is concerned with the structure, function, and regulation of cell-surface molecules that mediate recognition phenomena during embryogenesis. Area-code molecules are cell-surface molecules which are involved in the specific recognition phenomena during growth and development. These molecules provide cells with distinct cell-surface addresses or phenotypes, and provide the basis for the specificity in cell-cell recognition during cell migrations and cell-cell interactions, as well as serving as receptors for diffusible differentiation signals. The area-code hypothesis has 3 main postulates. i) There is a progressive display of specific combinations of area-code molecules on the surfaces of cells during development. ii) The genetic programs which determine the specific expression of area-code molecules are in part controlled by DNA modifications. These chromosomal modifications are believed to channel cells into specific lineages uith progressively restricted developmental options...

Animals

Maternal obesity in rats results in male-specific increases in genome-wide DNA methylation in postnatal offspring liver.

Male-specific peripubertal DNA demethylation in the liver has been reported in mice. Here, we investigated whether it also occurs in rats, the influence of maternal obesity and whether DNA demethylation changes contribute to observed sex-specific effects of maternal obesity in offspring. Female rats were fed a high-fat, high-sugar 'cafeteria' (Caf) diet before mating with standard chow-fed males. The offspring liver methylome and transcriptome were examined. Body weight was higher in Caf-fed dams prior to mating, during gestation and at parturition. Male and female offspring from Caf-fed dams had lower birth weights but higher adult weights and adiposity than offspring from chow-fed dams. A comparison of DNA methylation in 3-week-old weaner males versus female siblings from chow-fed dams did not reveal the male-specific DNA demethylation that was previously reported in mice. However, strong maternal diet effects in male weaner offspring methylation were observed. A comparison of female weaners from chow- versus Caf-fed dams showed a range of differences, with 39% of differentially methylated regions (DMRs) having higher methylation in Caf offspring and 61% of DMRs having higher methylation in chow offspring. In stark contrast, 99% of maternal-diet-induced DMRs in male weaner offspring had higher methylation in offspring from Caf-fed dams. This suggests that maternal obesity induces widespread hypermethylation in the male offspring liver at weaning. However, a comparison with RNA sequencing data revealed limited transcriptional changes at this developmental stage or in adult offspring. While these data highlight how environmentally sensitive DNA methylation is in the male rodent perinatal period, these methylation changes may not be a major contributor to sex differences in developmentally programmed liver disease.

Animals

Repurposing anti-phage defenses to differentially arrest the viral lifecycle reveals the regulatory logic of a parasitic satellite.

Mobile genetic elements frequently encode defense mechanisms to protect their bacterial hosts from viral attack. In Vibrio cholerae, these defensive elements include phage-inducible chromosomal island-like elements (PLEs), which are phage satellites that act as highly specialized parasites of the lytic phage ICP1. While PLE transcriptional activation upon ICP1 infection is known to be temporally regulated, the underlying regulatory logic and dependencies on the progression of the phage's developmental program required for activation remain unclear. In this study, we took a novel approach to define these dependencies by introducing independent anti-phage defense systems, BREX and DarTG, as molecular roadblocks to impede the ICP1 lifecycle. We discovered that, for both ICP1 and PLE, late-stage gene expression is fundamentally uncoupled from genome replication, representing a striking departure from the standard paradigm for double-stranded DNA phages. While BREX restricts ICP1 to an immediate-early transcriptional state that stalls PLE activation, DarTG allows the phage to execute its full transcriptional cascade despite the total block in DNA replication. This permissive environment provides the necessary cues for complete PLE induction, revealing that the extent of ICP1 transcriptional progression is a key determinant of PLE transcriptional activation. Unlike other phage satellites that rely on a single cue for activation, our results demonstrate that PLE uses a progressive licensing strategy that relies on multiple cues tied to milestones in the phage's developmental program. This regulatory architecture ensures robust PLE activation resilient to phage escape.

Journal Article

Allied health careers special resources and services program: increasing the probabilities of success for 'high risk' students in allied health career programs.

This article describes a successful developmental program specifically designed for academically "high risk" students entering a two-year community college career program in allied health. The program consisted of providing an intensive three-week instructional program to students before they entered the allied health career program, and subsequently providing an ongoing support system of tutoring, counseling and career development activities. Participants attained higher levels of academic performance and retention than nonparticipants.

Achievement

Gametogenic processes and their relationship to normal and abnormal conceptus development.

The relationship between gametogenesis and conceptus development has been described in relation to recent experiments concerning selected aspects of the cellular and molecular basis of differentiation of female and male gametes. Evidence suggests that a variety of hormonal and nonhormonal mechanisms or combinations thereof have evolved and control distinct stages of oocyte maturation process. Analyses of the meiotic maturation process with particular emphasis on amphibians, indicate that distinct cytoplasmic factors are produced or activated in response to a hormonal stimulus which initiates germinal vesicle breakdown and reinitiates the meiotic maturation process. Evidence suggests that certain cytoplasmic factors originate in the nucleus and (or) cytoplasm and that considerable cytoplasmic maturation proceeds in the absence of the nucleus, or as a result of nuclear-cytoplasmic interactions. Cytoplasmic factor(s) or activities affect many aspects of oocyte function and structure, including the cell membrane, chromosomes, nucleus and meiotic spindle. Ions also play a crucial role in gamete differentiation either alone or in combination with hormones or these cytoplasmic factors. The relevance of these findings to oocyte maturation and fertilization, activation and embryonic development is discussed. In many cases, similar types of biological activities exist in widely separated species and (or) are effective in widely different species. The importance of synchronization of normal gametes to the development of the embryos is discussed. It is proposed, and the evidence suggests, that hormones, ions and cytoplasmic factors play a fundamental and variable role in the differentiation and function of "fully" mature oocytes of many species including mammals. Significantly, the data suggest that a "fully" mature oocyte occurs as a result of the expression of an underlying developmental program. Variations in these substances or the processes involved in their formation or actions appear to be relevant to understanding a wide variety of developmental abnormalities as well as to assessing the normality or abnormality of in vivo or in vitro maturation and differentiation.

Animals

A conserved partner-switching system controls terminal differentiation in multicellular cyanobacteria.

UNLABELLED: Canonical partner-switching systems (PSSs) regulate sigma factor activity through reversible phosphorylation, but their established roles have been largely limited to stress responses and sporulation in Firmicutes. Whether this regulatory mechanism also controls developmental cell fate decisions in other bacterial phyla has remained unknown. Here, we identify a canonical PSS that governs heterocyst differentiation in the multicellular cyanobacterium Anabaena sp. PCC 7120. This system comprises the anti-sigma factor All2284 (NfsS) and the anti-anti-sigma factor All2283 (NfsR). Structural predictions and biochemical assays showed that NfsS phosphorylates NfsR on a conserved serine residue, whereas bacterial two-hybrid and co-purification assays demonstrated that NfsS binds the developmental sigma factors SigC and SigE. Deletion of nfsR abolished heterocyst formation and diazotrophic growth, and transcriptomic analysis revealed broad failure to induce late heterocyst genes, including nitrogen fixation functions such as nifHDK and fdxH. Phylum-wide comparative genomics further showed that PSS genes and putative functional clusters are strongly enriched in filamentous and heterocyst-forming taxa, indicating an association between the expansion of these signaling modules and the emergence of multicellularity and developmental specialization. Together, these findings establish a PSS as a direct regulator of terminal cell differentiation in a gram-negative bacterium and reveal partner switching as a conserved regulatory principle linking environmental signaling to developmental fate in a major bacterial phylum. IMPORTANCE: While partner-switching systems are classically associated with stress responses and sporulation control in Firmicutes, whether this regulatory logic governs developmental decisions in other bacterial phyla has remained unknown. Here, we establish that a related partner-switching mechanism operates in a distinct bacterial lineage, the cyanobacteria, where it controls a major developmental transition involving terminal cell differentiation. Specifically, we show that a phosphorylation-dependent checkpoint involving the anti-sigma factor NfsS and the anti-anti-sigma factor NfsR directly regulates heterocyst formation. Disruption of this switch abolishes cell differentiation and diazotrophic growth, revealing that this system is an obligate gatekeeper for terminal differentiation. Conceptually, these findings substantially extend the known functional repertoire of partner-switching circuits: rather than controlling stress adaptation or spore dormancy, this module has been co-opted to govern a complex, multicellular developmental program in an organism that underpins global carbon and nitrogen cycles. This work, therefore, establishes a new paradigm for phosphorylation-based control of developmental sigma factors and provides a tractable model for dissecting how conserved signaling modules are rewired to drive lineage-specific innovations across the bacterial domain.

cell differentiation