A phosphorylation code regulates the multi-functional protein RETINOBLASTOMA-RELATED1 in Arabidopsis thaliana

The RETINOBLASTOMA-RELATED (RBR) proteins play a central role coordinating cell division, cell differentiation and cell survival within an environmental and developmental context. These roles reflect RBR ability to engage in multiple protein-protein interactions (PPIs), which are regulated by multi-site phosphorylation. However the functional outcomes of RBR phosphorylation in multicellular organisms remain largely unexplored. Here we test the hypothesis that phosphorylation allows diversification of RBR functions in multicellular context. Using a representative collection of transgenic loss- and gain of function point mutations in RBR phospho-sites, we analysed their complementation capacity in Arabidopsis thaliana root meristems. While the number of mutated residues often correlated to the phenotypic strength of RBR phospho-variants, phospho-sites contributed differentially to distinct phenotypes. For example, the pocket-domain has a greater influence on meristematic cell proliferation, whereas the C-terminal region associates to stem cell maintenance. We found combinatorial effects between the T406 phopspho-site with others in different protein domains. Moreover, a phospho-mimetic and a phospho-defective variant, both promoting cell death, indicate that RBR controls similar cell fate choices by distinct mechanisms. Thus, additivity and specificity of RBR phospho-sites fine tune RBR activity across its multiple roles. Interestingly, a mutation disrupting RBR interactions with the LXCXE motif suppresses dominant phospho-defective RBR phenotypes. By probing protein-protein interactions of RBR variants, we found that LXCXE-containing members of the DREAM complex constitute an important component of phosphorylation-regulated RBR function, but also that RBR participates in stress or environmental responses independently of its phosphorylation state. We conclude that developmental-related, but not stress- or environmental-related functions of RBR are defined and separable by a combinatorial phosphorylation code.

Ontogeny of the Projections From the Dorsomedial Division of the Anterior Bed Nucleus of the Stria Terminalis to Hypothalamic Nuclei

The bed nucleus of the stria terminalis (BNST) is a telencephalic structure well-connected to hypothalamic regions known to control goal-oriented behaviors such as feeding. In particular, we showed that the dorsomedial division of the anterior BNST innervate neurons of the paraventricular (PVH), dorsomedial (DMH), and arcuate (ARH) hypothalamic nuclei as well as the lateral hypothalamic area (LHA). While the anatomy of these projections has been characterized in mice, their ontogeny has not been studied. In this study, we used the DiI-based tract tracing approach to study the development of BNST projections innervating several hypothalamic areas including the PVH, DMH, ARH, and LHA. These results indicate that projections from the dorsomedial division of the anterior BNST to hypothalamic nuclei are immature at birth and substantially reach the PVH, DMH, and the LHA at P10. In the ARH, only sparse fibers are observed at P10, but their density increased markedly between P12 and P14. Collectively, these findings provide new insight into the ontogeny of hypothalamic circuits, and highlight the importance of considering the developmental context as a direct modulator in their proper formation.

Notch-dependent DNA cis-regulatory elements and their dose-dependent control of C. elegans stem cell self-renewal

A long-standing biological question is how DNA cis-regulatory elements shape transcriptional patterns during metazoan development. The use of reporter constructs, cell culture and computational modeling has made enormous contributions to understanding this fundamental question, but analysis of regulatory elements in their natural developmental context is an essential but rarely used complement. Here, we edited Notch-dependent cis-regulatory elements in the endogenous C. elegans sygl-1 gene, which encodes a key stem cell regulator. We then analyzed the in vivo consequences of those mutations – on both gene expression (nascent transcripts, mRNA, protein) and stem cell maintenance. Mutation of a single element in a three-element homotypic cluster reduced expression as well as stem cell pool size by about half, while mutation of two elements essentially abolished them. We find that LBS number and LBS neighborhood are both important to activity: elements on separate chromosomes function additively, while elements in the same cluster act synergistically. Our approach of precise CRISPR/Cas9 gene editing coupled with quantitation of both molecular and biological readouts establishes a powerful model for in vivo functional analyses of DNA cis-regulatory elements.

Alveologenesis: What Governs Secondary Septa Formation

The simplification of alveoli leads to various lung pathologies such as bronchopulmonary dysplasia and emphysema. Deep insight into the process of emergence of the secondary septa during development and regeneration after pneumonectomy, and into the contribution of the drivers of alveologenesis and neo-alveolarization is required in an efficient search for therapeutic approaches. In this review, we describe the formation of the gas exchange units of the lung as a multifactorial process, which includes changes in the actomyosin cytoskeleton of alveocytes and myofibroblasts, elastogenesis, retinoic acid signaling, and the contribution of alveolar mesenchymal cells in secondary septation. Knowledge of the mechanistic context of alveologenesis remains incomplete. The characterization of the mechanisms that govern the emergence and depletion of αSMA will allow for an understanding of how the niche of fibroblasts is changing. Taking into account the intense studies that have been performed on the pool of lung mesenchymal cells, we present data on the typing of interstitial fibroblasts and their role in the formation and maintenance of alveoli. On the whole, when identifying cell subpopulations in lung mesenchyme, one has to consider the developmental context, the changing cellular functions, and the lability of gene signatures.

Dissecting CTCF site function in a tense HoxD locus

In this issue of Genes & Development, Amândio and colleagues (pp. 1490–1509) dissect the function of a cluster of several CTCF binding sites in the HoxD cluster by iterative deletions in mice. They found additive functions for some, and intriguingly found that some sites function as insulators, while others function as anchors for enhancer–promoter interactions. These functions vary depending on developmental context. The work provides new insights into the roles played by CTCF in regulating developmental patterns and 3D chromatin organization.

Supporting the development of children with moderate and severe intellectual disabilities in a special kindergarten – a practical approach

The aim of this article is to characterize the cognitive development of a preschool child on the basis of selected theoretical concepts in relation to moderate and severe intellectual disability. These considerations will serve as a guideline for further theoretical analyses and practical activities supporting the development of children with intellectual disabilities. From a biological perspective, intellectual disability is a typical medical problem. In the developmental context, it means “a failure in the process of development” (Kościelska 1995; Speck, 2015). I assume that the determinant for planning and implementing the content, forms and methods of teaching in special kindergarten should be the level of cognitive development of each child. In the study I refer to selected concepts of development in the terms of Jean Piaget and Lev Vygotsky, which, as I assume, can be a field for analysis of the development of children who require multi-profile support.

Expression of human HIPKs in Drosophila demonstrates their shared and unique functions in a developmental model

Homeodomain-interacting protein kinases (HIPKs) are a family of four conserved proteins essential for vertebrate development, as demonstrated by defects in the eye, brain, and skeleton that culminate in embryonic lethality when multiple HIPKs are lost in mice. While HIPKs are essential for development, functional redundancy between the four vertebrate HIPK paralogues has made it difficult to compare their respective functions. Because understanding the unique and shared functions of these essential proteins could directly benefit the fields of biology and medicine, we addressed the gap in knowledge of the four vertebrate HIPK paralogues by studying them in the fruit fly Drosophila melanogaster, where reduced genetic redundancy simplifies our functional assessment. The single hipk present in the fly allowed us to perform rescue experiments with human HIPK genes that provide new insight into their individual functions not easily assessed in vertebrate models. Further, the abundance of genetic tools and established methods for monitoring specific developmental pathways and gross morphological changes in the fly allowed for functional comparisons in endogenous contexts. We first performed rescue experiments to demonstrate the extent to which each of the human HIPKs can functionally replace Drosophila Hipk for survival and morphological development. We then showed the ability of each human HIPK to modulate Armadillo/β-catenin levels, JAK/STAT activity, proliferation, growth, and death, each of which have previously been described for Hipks, but never all together in comparable tissue contexts. Finally, we characterized novel developmental phenotypes induced by human HIPKs to gain insight to their unique functions. Together, these experiments provide the first direct comparison of all four vertebrate HIPKs to determine their roles in a developmental context.

Early Childhood Obesity: A Developmental Perspective

Childhood obesity is a multifactorial disease, shaped by child, familial, and societal influences; prevention efforts must begin early in childhood. Viewing the problem of childhood obesity through a developmental lens is critical to understanding the nuances of a child's interactions with food and their environment across the span of growth and development. Risk factors for childhood obesity begin prior to birth, compounding across the life course. Some significant risk factors are unmodifiable (e.g., genetics) while others are theoretically modifiable. Social inequities, however, hinder many families from easily making modifications to a range of risk factors. The objective of this review is to provide background and an overview of the literature on childhood obesity in early childhood (birth to 5 years of age) in a developmental context. Special focus is placed on unique developmental considerations, child eating behaviors, and parental feeding behaviors in infancy, toddlerhood, and preschool ages. Expected final online publication date for the Annual Review of Developmental Psychology, Volume 3 is December 2021. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.