M1BP is an essential transcriptional activator of oxidative metabolism during Drosophila development.

Oxidative metabolism is the predominant energy source for aerobic muscle contraction in adult animals. How the cellular and molecular components that support aerobic muscle physiology are put in place during development through their transcriptional regulation is not well understood. Using the Drosophila flight muscle model, we show that the formation of mitochondria cristae harbouring the respiratory chain is concomitant with a large-scale transcriptional upregulation of genes linked with oxidative phosphorylation (OXPHOS) during specific stages of flight muscle development. We further demonstrate using high-resolution imaging, transcriptomic and biochemical analyses that Motif-1-binding protein (M1BP) transcriptionally regulates the expression of genes encoding critical components for OXPHOS complex assembly and integrity. In the absence of M1BP function, the quantity of assembled mitochondrial respiratory complexes is reduced and OXPHOS proteins aggregate in the mitochondrial matrix, triggering a strong protein quality control response. This results in isolation of the aggregate from the rest of the matrix by multiple layers of the inner mitochondrial membrane, representing a previously undocumented mitochondrial stress response mechanism. Together, this study provides mechanistic insight into the transcriptional regulation of oxidative metabolism during Drosophila development and identifies M1BP as a critical player in this process.

Sustained experimental activation of FGF8/ERK in the developing chicken spinal cord models early events in ERK-mediated tumorigenesis.

The MAPK/ERK pathway regulates a variety of physiological cellular functions, including cell proliferation and survival. It is abnormally activated in many types of human cancers in response to driver mutations in regulators of this pathway that trigger tumor initiation. The early steps of oncogenic progression downstream of ERK overactivation are poorly understood due to a lack of appropriate models. We show here that ERK1/2 overactivation in the trunk neural tube of the chicken embryo through expression of a constitutively active form of the upstream kinase MEK1 (MEK1ca), rapidly provokes a profound change in the transcriptional signature of developing spinal cord cells. These changes are concordant with a previously established role of the tyrosine kinase receptor ligand FGF8 acting via the ERK1/2 effectors to maintain an undifferentiated state. Furthermore, we show that MEK1ca-transfected spinal cord cells lose neuronal identity, retain caudal markers, and ectopically express potential effector oncogenes, such as AQP1. MEK1ca expression in the developing spinal cord from the chicken embryo is thus a tractable in vivo model to identify the mechanisms fostering neoplasia and malignancy in ERK-induced tumorigenesis of neural origins.

Hox Proteins in the Regulation of Muscle Development.

Hox genes encode evolutionary conserved transcription factors that specify the anterior-posterior axis in all bilaterians. Being well known for their role in patterning ectoderm-derivatives, such as CNS and spinal cord, Hox protein function is also crucial in mesodermal patterning. While well described in the case of the vertebrate skeleton, much less is known about Hox functions in the development of different muscle types. In contrast to vertebrates however, studies in the fruit fly, Drosophila melanogaster, have provided precious insights into the requirement of Hox at multiple stages of the myogenic process. Here, we provide a comprehensive overview of Hox protein function in Drosophila and vertebrate muscle development, with a focus on the molecular mechanisms underlying target gene regulation in this process. Emphasizing a tight ectoderm/mesoderm cross talk for proper locomotion, we discuss shared principles between CNS and muscle lineage specification and the emerging role of Hox in neuromuscular circuit establishment.

HOXB8 Counteracts MAPK/ERK Oncogenic Signaling in a Chicken Embryo Model of Neoplasia.

HOX transcription factors are members of an evolutionarily conserved family of proteins required for the establishment of the anteroposterior body axis during bilaterian development. Although they are often deregulated in cancers, the molecular mechanisms by which they act as oncogenes or tumor suppressor genes are only partially understood. Since the MAPK/ERK signaling pathway is deregulated in most cancers, we aimed at apprehending if and how the Hox proteins interact with ERK oncogenicity. Using an in vivo neoplasia model in the chicken embryo consisting in the overactivation of the ERK1/2 kinases in the trunk neural tube, we analyzed the consequences of the HOXB8 gain of function at the morphological and transcriptional levels. We found that HOXB8 acts as a tumor suppressor, counteracting ERK-induced neoplasia. The HOXB8 tumor suppressor function relies on a large reversion of the oncogenic transcriptome induced by ERK. In addition to showing that the HOXB8 protein controls the transcriptional responsiveness to ERK oncogenic signaling, our study identified new downstream targets of ERK oncogenic activation in an in vivo context that could provide clues for therapeutic strategies.

HoxB genes regulate neuronal delamination in the trunk neural tube by controlling the expression of Lzts1.

Differential Hox gene expression is central for specification of axial neuronal diversity in the spinal cord. Here, we uncover an additional function of Hox proteins in the developing spinal cord, restricted to B cluster Hox genes. We found that members of the HoxB cluster are expressed in the trunk neural tube of chicken embryo earlier than Hox from the other clusters, with poor antero-posterior axial specificity and with overlapping expression in the intermediate zone (IZ). Gain-of-function experiments of HoxB4, HoxB8 and HoxB9, respectively, representative of anterior, central and posterior HoxB genes, resulted in ectopic progenitor cells in the mantle zone. The search for HoxB8 downstream targets in the early neural tube identified the leucine zipper tumor suppressor 1 gene (Lzts1), the expression of which is also activated by HoxB4 and HoxB9. Gain- and loss-of-function experiments showed that Lzts1, which is expressed endogenously in the IZ, controls neuronal delamination. These data collectively indicate that HoxB genes have a generic function in the developing spinal cord, controlling the expression of Lzts1 and neuronal delamination.

Human ZKSCAN3 and Drosophila M1BP are functionally homologous transcription factors in autophagy regulation.

Autophagy is an essential cellular process that maintains homeostasis by recycling damaged organelles and nutrients during development and cellular stress. ZKSCAN3 is the sole identified master transcriptional repressor of autophagy in human cell lines. How ZKSCAN3 achieves autophagy repression at the mechanistic or organismal level however still remains to be elucidated. Furthermore, Zkscan3 knockout mice display no discernable autophagy-related phenotypes, suggesting that there may be substantial differences in the regulation of autophagy between normal tissues and tumor cell lines. Here, we demonstrate that vertebrate ZKSCAN3 and Drosophila M1BP are functionally homologous transcription factors in autophagy repression. Expression of ZKSCAN3 in Drosophila prevents premature autophagy onset due to loss of M1BP function and conversely, M1BP expression in human cells can prevent starvation-induced autophagy due to loss of nuclear ZKSCAN3 function. In Drosophila ZKSCAN3 binds genome-wide to sequences targeted by M1BP and transcriptionally regulates the majority of M1BP-controlled genes, demonstrating the evolutionary conservation of the transcriptional repression of autophagy. This study thus  allows the potential for transitioning the mechanisms, gene targets and plethora metabolic processes controlled by M1BP onto ZKSCAN3 and opens up Drosophila as a tool in studying the function of ZKSCAN3 in autophagy and tumourigenesis.

Cooperation of axial and sex specific information controls Drosophila female genitalia growth by regulating the Decapentaplegic pathway.

The specification and morphogenesis of an organ requires the coordinate deployment and integration of regulatory information, including sex specific information when the organ is sex specific. Only a few gene networks controlling size and pattern development have been deciphered, which limits the emergence of principles, general or not, underlying the organ-specifying gene networks. Here we elucidate the genetic and molecular network determining the control of size in the Drosophila abdominal A9 primordium, contributing to the female genitalia. This network requires axial regulatory information provided by the Hox protein Abdominal-BR (Abd-BR), the Hox cofactors Extradenticle (Exd) and Homothorax (Hth), and the sex specific transcription factor Doublesex Female (DsxF). These factors synergize to control size in the female A9 by the coordinate regulation of the Decapentaplegic (Dpp) growth pathway. Molecular dissection of the dpp regulatory region and in vivo protein interaction experiments suggest that Abd-BR, Exd, Hth and DsxF coordinately regulate a short dpp enhancer to repress dpp expression and restrict female A9 size. The same regulators can also suppress dpp expression in the A8, but this requires the absence of the Abd-BM isoform, which specifies A8. These results delineate the network controlling female A9 growth in Drosophila.

The Generic Facet of Hox Protein Function.

Hox transcription factors are essential to promote morphological diversification of the animal body. A substantial number of studies have focused on how Hox proteins reach functional specificity, an issue that arises from the fact that these transcription factors control distinct developmental functions despite sharing similar molecular properties. In this review, we highlight that, besides specific functions, for which these transcription factors are renowned, Hox proteins also often have nonspecific functions. We next discuss some emerging principles of these generic functions and how they relate to specific functions and explore our current grasp of the underlying molecular mechanisms.

Post-translational modifications of HOX proteins, an underestimated issue.

Post-translational modifications (PTMs) are important determinants which contribute to modulating the turn-over, intracellular localisation, molecular interactions and activity of most eukaryotic proteins. Such modifications and their consequences have been extensively examined for some proteins or classes of proteins. This is not the case for the HOX transcription factors which are crucial regulators of animal development. In this review, we provide a survey of the literature and data repositories pertaining to HOX-associated PTMs. This highlights that HOX proteins are also likely widely post-translationally modified, and defines HOX PTMs as an under-valued facet of their biology.

Fattening the perspective of Hox protein specificity through SLiMming.

The functional identification and dissection of protein domains has been a successful approach towards the understanding of Hox protein specificity. However, only a few functional protein domains have been identified; this has been a major limitation in deciphering the molecular modalities of Hox protein action. We explore here, by in silico survey of short linear motifs (SLiMs) in Hox proteins, the contribution of SLiMs to Hox proteins, focusing on the mouse, chick and Drosophila Hox complement. Our findings reveal a widespread and uniform distribution of SLiMs along Hox protein sequences and identify the most apparent features of Hox associated SLiMs. While few motifs have been associated with Hox proteins so far, this work suggests that many more contribute to Hox protein functions. The potential and difficulties to apprehend the full contribution of SLiMs in controlling Hox protein functions are discussed.

The Hox proteins Ubx and AbdA collaborate with the transcription pausing factor M1BP to regulate gene transcription.

In metazoans, the pausing of RNA polymerase II at the promoter (paused Pol II) has emerged as a widespread and conserved mechanism in the regulation of gene transcription. While critical in recruiting Pol II to the promoter, the role transcription factors play in transitioning paused Pol II into productive Pol II is, however, little known. By studying how Drosophila Hox transcription factors control transcription, we uncovered a molecular mechanism that increases productive transcription. We found that the Hox proteins AbdA and Ubx target gene promoters previously bound by the transcription pausing factor M1BP, containing paused Pol II and enriched with promoter-proximal Polycomb Group (PcG) proteins, yet lacking the classical H3K27me3 PcG signature. We found that AbdA binding to M1BP-regulated genes results in reduction in PcG binding, the release of paused Pol II, increases in promoter H3K4me3 histone marks and increased gene transcription. Linking transcription factors, PcG proteins and paused Pol II states, these data identify a two-step mechanism of Hox-driven transcription, with M1BP binding leading to Pol II recruitment followed by AbdA targeting, which results in a change in the chromatin landscape and enhanced transcription.

Hox functional diversity: Novel insights from flexible motif folding and plastic protein interaction.

How the formidable diversity of forms emerges from developmental and evolutionary processes is one of the most fascinating questions in biology. The homeodomain-containing Hox proteins were recognized early on as major actors in diversifying animal body plans. The molecular mechanisms underlying how this transcription factor family controls a large array of context- and cell-specific biological functions is, however, still poorly understood. Clues to functional diversity have emerged from studies exploring how Hox protein activity is controlled through interactions with PBC class proteins, also evolutionary conserved HD-containing proteins. Recent structural data and molecular dynamic simulations add further mechanistic insights into Hox protein mode of action, suggesting that flexible folding of protein motifs allows for plastic protein interaction. As we discuss in this review, these findings define a novel type of Hox-PBC interaction, weak and dynamic instead of strong and static, hence providing novel clues to understanding Hox transcriptional specificity and diversity.

Hox Service Warranty Extends to Adult Bone Repair.

Hox genes are key developmental regulators. In this issue of Developmental Cell, Rux et al. (2016) uncover an adult role for Hox11 genes in regionalized bone repair. This function relies on Hox activity in bone marrow multipotent mesenchymal stem progenitor cells, which promotes skeletal cell differentiation.

Autophagy : Moving Benchside Promises to Patient Bedsides.

Survival rates of patients with metastatic or recurrent cancers have remained virtually unchanged during the past 30 years. This fact makes the need for new therapeutic options even more urgent. An attractive option would be to target autophagy, an essential quality control process that degrades toxic aggregates, damaged organelles, and signaling proteins, and acts as a tumor suppressor pathway of tumor initiation. Conversely, other fascinating observations suggest that autophagy supports cancer progression, relapse, metastasis, dormancy and resistance to therapy. This review provides an overview of the contradictory roles that autophagy plays in cancer initiation and progression and discusses the promises and challenges of current strategies that target autophagy for cancer therapy.

A survey of conservation of sea spider and Drosophila Hox protein activities.

Hox proteins have well-established functions in development and evolution, controlling the final morphology of bilaterian animals. The common phylogenetic origin of Hox proteins and the associated evolutionary diversification of protein sequences provide a unique framework to explore the relationship between changes in protein sequence and function. In this study, we aimed at questioning how sequence variation within arthropod Hox proteins influences function. This was achieved by exploring the functional impact of sequence conservation/divergence of the Hox genes, labial, Sex comb reduced, Deformed, Ultrabithorax and abdominalA from two distant arthropods, the sea spider and the well-studied Drosophila. Results highlight a correlation between sequence conservation within the homeodomain and the degree of functional conservation, and identify a novel functional domain in the Labial protein.

Cellular and molecular insights into Hox protein action.

Hox genes encode homeodomain transcription factors that control morphogenesis and have established functions in development and evolution. Hox proteins have remained enigmatic with regard to the molecular mechanisms that endow them with specific and diverse functions, and to the cellular functions that they control. Here, we review recent examples of Hox-controlled cellular functions that highlight their versatile and highly context-dependent activity. This provides the setting to discuss how Hox proteins control morphogenesis and organogenesis. We then summarise the molecular modalities underlying Hox protein function, in particular in light of current models of transcription factor function. Finally, we discuss how functional divergence between Hox proteins might be achieved to give rise to the many facets of their action.

A flexible extension of the Drosophila ultrabithorax homeodomain defines a novel Hox/PBC interaction mode.

The patterning function of Hox proteins relies on assembling protein complexes with PBC proteins, which often involves a protein motif found in most Hox proteins, the so-called Hexapeptide (HX). Hox/PBC complexes likely gained functional diversity by acquiring additional modes of interaction. Here, we structurally characterize the first HX alternative interaction mode based on the paralogue-specific UbdA motif and further functionally validate structure-based predictions. The UbdA motif folds as a flexible extension of the homeodomain recognition helix and defines Hox/PBC contacts that occur, compared with those mediated by the HX motif, on the opposing side of the DNA double helix. This provides a new molecular facet to Hox/PBC complex assembly and suggests possible mechanisms for the diversification of Hox protein function.

Drosophila melanogaster Hox transcription factors access the RNA polymerase II machinery through direct homeodomain binding to a conserved motif of mediator subunit Med19.

Hox genes in species across the metazoa encode transcription factors (TFs) containing highly-conserved homeodomains that bind target DNA sequences to regulate batteries of developmental target genes. DNA-bound Hox proteins, together with other TF partners, induce an appropriate transcriptional response by RNA Polymerase II (PolII) and its associated general transcription factors. How the evolutionarily conserved Hox TFs interface with this general machinery to generate finely regulated transcriptional responses remains obscure. One major component of the PolII machinery, the Mediator (MED) transcription complex, is composed of roughly 30 protein subunits organized in modules that bridge the PolII enzyme to DNA-bound TFs. Here, we investigate the physical and functional interplay between Drosophila melanogaster Hox developmental TFs and MED complex proteins. We find that the Med19 subunit directly binds Hox homeodomains, in vitro and in vivo. Loss-of-function Med19 mutations act as dose-sensitive genetic modifiers that synergistically modulate Hox-directed developmental outcomes. Using clonal analysis, we identify a role for Med19 in Hox-dependent target gene activation. We identify a conserved, animal-specific motif that is required for Med19 homeodomain binding, and for activation of a specific Ultrabithorax target. These results provide the first direct molecular link between Hox homeodomain proteins and the general PolII machinery. They support a role for Med19 as a PolII holoenzyme-embedded "co-factor" that acts together with Hox proteins through their homeodomains in regulated developmental transcription.

Hox proteins mediate developmental and environmental control of autophagy.

Hox genes encode evolutionarily conserved transcription factors, providing positional information used for differential morphogenesis along the anteroposterior axis. Here, we show that Drosophila Hox proteins are potent repressors of the autophagic process. In inhibiting autophagy, Hox proteins display no apparent paralog specificity and do not provide positional information. Instead, they impose temporality on developmental autophagy and act as effectors of environmental signals in starvation-induced autophagy. Further characterization establishes that temporality is controlled by Pontin, a facultative component of the Brahma chromatin remodeling complex, and that Hox proteins impact on autophagy by repressing the expression of core components of the autophagy machinery. Finally, the potential of central and posterior mouse Hox proteins to inhibit autophagy in Drosophila and in vertebrate COS-7 cells indicates that regulation of autophagy is an evolutionary conserved feature of Hox proteins.