Posterior Wnts have distinct roles in specification and patterning of the planarian posterior region

The wnt signaling pathway is an intercellular communication mechanism essential in cell-fate specification, tissue patterning and regional-identity specification. A βcatenin-dependent signal specifies the AP (Anteroposterior) axis of planarians, both during regeneration of new tissues and during normal homeostasis. Accordingly, four wnts (posterior wnts) are expressed in a nested manner in central and posterior regions of planarians. We have analyzed the specific role of each posterior wnt and the possible cooperation between them in specifying and patterning planarian central and posterior regions. We show that each posterior wnt exerts a distinct role during re-specification and maintenance of the central and posterior planarian regions, and that the integration of the different wnt signals (βcatenin dependent and independent) underlies the patterning of the AP axis from the central region to the tip of the tail. Based on these findings and data from the literature, we propose a model for patterning the planarian AP axis

The pioneer factor Smed-gata456-1 is required for gut cell differentiation and maintenance in planarians

How adult stem cells differentiate into different cell types remains one of the most intriguing questions in regenerative medicine. Pioneer factors are transcription factors that can bind to and open chromatin, and are among the first elements involved in cell differentiation. We used the freshwater planarian Schmidtea mediterranea as a model system to study the role of the gata456 family of pioneer factors in gut cell differentiation during both regeneration and maintenance of the digestive system. Our findings reveal the presence of two members of the gata456 family in the Schmidtea mediterranea genome; Smed-gata456-1 and Smed-gata456-2. Our results show that Smed-gata456-1 is the only ortholog with a gut cell-related function. Smed-gata456-1 is essential for the differentiation of precursors into intestinal cells and for the survival of these differentiated cells, indicating a key role in gut regeneration and maintenance. Furthermore, tissues other than the gut appear normal following Smed-gata456-1 RNA interference (RNAi), indicating a gut-specific function. Importantly, different neoblast subtypes are unaffected by Smed-gata456-1(RNAi), suggesting that 1) Smed-gata456-1 is involved in the differentiation and maintenance, but not in the early determination, of gut cells; and 2) that the stem cell compartment is not dependent on a functional gut

A C-terminally truncated form of β-catenin acts as a novel regulator of Wnt/β-catenin signaling in planarians

β-Catenin, the core element of the Wnt/β-catenin pathway, is a multifunctional and evolutionarily conserved protein which performs essential roles in a variety of developmental and homeostatic processes. Despite its crucial roles, the mechanisms that control its context-specific functions in time and space remain largely unknown. The Wnt/β-catenin pathway has been extensively studied in planarians, flatworms with the ability to regenerate and remodel the whole body, providing a 'whole animal' developmental framework to approach this question. Here we identify a C-terminally truncated β-catenin (β-catenin4), generated by gene duplication, that is required for planarian photoreceptor cell specification. Our results indicate that the role of β-catenin4 is to modulate the activity of β-catenin1, the planarian β-catenin involved in Wnt signal transduction in the nucleus, mediated by the transcription factor TCF-2. This inhibitory form of β-catenin, expressed in specific cell types, would provide a novel mechanism to modulate nuclear β-catenin signaling levels. Genomic searches and in vitro analysis suggest that the existence of a C-terminally truncated form of β-catenin could be an evolutionarily conserved mechanism to achieve a fine-tuned regulation of Wnt/β-catenin signaling in specific cellular contexts

Analysis of Fox genes in Schmidtea mediterranea reveals new families and a conserved role of Smed‑foxO in controlling cell death

The forkhead box (Fox) genes encode transcription factors that control several key aspects of development. Present in the ancestor of all eukaryotes, Fox genes underwent several duplications followed by loss and diversification events that gave rise to the current 25 families. However, few Fox members have been identified from the Lophotrochozoa clade, and specifically from planarians, which are a unique model for understanding development, due to the striking plasticity of the adult. The aim of this study was to identify and perform evolutionary and functional studies of the Fox genes of lophotrochozoan species and, specifically, of the planarian Schmidtea mediterranea. Generating a pipeline for identifying Forkhead domains and using phylogenetics allowed us the phylogenetic reconstruction of Fox genes. We corrected the annotation for misannotated genes and uncovered a new family, the QD, present in all metazoans. According to the new phylogeny, the 27 Fox genes found in Schmidtea mediterranea were classified into 12 families. In Platyhelminthes, family losses were accompanied by extensive gene diversification and the appearance of specific families, the A(P) and N(P). Among the newly identified planarian Fox genes, we found a single copy of foxO, which shows an evolutionary conserved role in controlling cell death

La regeneració i l'homeòstasi en les planàries, un model clàssic de biologia del desenvolupament

La regeneració és la capacitat d'un organisme de reemplaçar fragments perduts a causa d'una amputació traumàtica o degeneració. La regeneració de les noves estructures té lloc bé a partir de proliferació cel·lular i formació de novo, bé per remodelació dels teixits preexistents. Les planàries poden regenerar un nou organisme sencer a partir de petits fragments del seu cos. Aquest fet ha atret l'interès dels científics al llarg de la història. El 1814, Dalyell conclou que les planàries «es poden considerar immortals sota la fulla d'una navalla». La regeneració en les planàries requereix la generació de teixit nou en el lloc de la ferida mitjançant proliferació cel·lular, que produeix un teixit nou indiferenciat, el blastema, i el remodelatge dels teixits preexistents per recuperar les proporcions del nou organisme regenerat. Una altra propietat espectacular de les planàries és la capacitat de créixer i decréixer segons la ingesta d'aliment. En tot moment, però, al llarg d'aquest creixement/decreixement es mantenen les proporcions corporals i funcions correctes, gràcies al control homeostàtic. Tota aquesta plasticitat és deguda, a escala cel·lular, a la presència de cèl·lules mare totipotents en un alt percentatge (entre el 20-30 % del total cel·lular en un organisme adult). Una altra propietat fonamental és la contínua activitat dels mecanismes morfogenètics, que normalment apareixen una sola vegada en el desenvolupament de la resta dels altres organismes. L'aplicació de noves metodologies a escala cel·lular, molecular i genètica en l'era postgenòmica ens ha permès estudiar funcionalment vies i gens del desenvolupament en un nou escenari, la regeneració de planàries

WNT-FRIZZLED-LRP5/6 Signaling Mediates Posterior Fate and Prolifer-ation during Planarian Regeneration

An organizer is defined as a group of cells that secrete extracellular proteins that specify the fate of surrounding cells according to their concentration. Their function during embryogenesis is key in patterning new growing tissues. Although organizers should also participate in adult development when new structures are regenerated, their presence in adults has only been identified in a few species with striking regenerative abilities, such as planarians. Planarians provide a unique model to understand the function of adult organizers, since the presence of adult pluripotent stem cells provides them with the ability to regenerate any body part. Previous studies have shown that the differential activation of the WNT/ -catenin signal in each wound is fundamental to establish an anterior or a posterior organizer in the corresponding wound. Here, we identify the receptors that mediate the WNT/ -catenin signal in posterior-facing wounds. We found that Wnt1-Fzd1-LRP5/6 signaling is evolutionarily conserved in executing a WNT/ -catenin signal to specify cell fate and to trigger a proliferative response. Our data allow a better understanding of the mechanism through which organizers signal to a "competent" field of cells and integrate the patterning and growth required during de novo formation of organs and tissues

FoxK1 is required for ectodermal cell differentiation during planarian regeneration

Forkhead box (Fox) genes belong to the 'winged helix' transcription factor superfamily. The function of some Fox genes is well known, such as the role of foxO in controlling metabolism and longevity and foxA in controlling differentiation of endodermal tissues. However, the role of some Fox factors is not yet well characterized. Such is the case of FoxK genes, which are mainly studied in mammals and have been implicated in diverse processes including cell proliferation, tissue differentiation and carcinogenesis. Planarians are free-living flatworms, whose importance in biomedical research lies in their regeneration capacity. Planarians possess a wide population of pluripotent adult stem cells, called neoblasts, which allow them to regenerate any body part after injury. In a recent study, we identified three foxK paralogs in the genome of Schmidtea mediterranea. In this study, we demonstrate that foxK1 inhibition prevents regeneration of the ectodermal tissues, including the nervous system and the epidermis. These results correlate with foxK1 expression in neoblasts and in neural progenitors. Although the triggering of wound genes expression, polarity reestablishment and proliferation was not affected after foxK1 silencing, the apoptotic response was decreased. Altogether, these results suggest that foxK1 would be required for differentiation and maintenance of ectodermal tissues

Descritpion of a double mutant strain of Drosophila melanogaster useful for genetic laboratory courses.

Many years ago, individuals showing drastically reduced eyes arose in our laboratory e (ebony) strain (Bridges and Morgan, 1923). We selected those flies presenting both traits and constituted a new double mutant strain e su (e, ebony; su, 'sense ulls', eyes drastically reduced). Both mutations were linked and located in the chromosome III. We used this strain in linkage analyses with our undergraduate students. [...

A hands-on genetics teaching approach at university level.

Teaching general Genetics is a cornerstone of a large number of university degrees. Being a scientific topic, laboratory classes are an essential element in student-centered learning. Here, we present our experience in implementing new material for teaching hands-on genetics, a subject of interest for other academic professionals in the field of Genetics. Our students carry out a genetic analysis of the su (sense ulls) mutation of Drosophila melanogaster, which produces a drastic eye reduction. The complete strain description can be found in Mestres et al. (2016a). The aim of the course is to give students the appropriate genetics tools to answer the three following questions: 1) Is the su mutation dominant or recessive? 2) In which chromosome is su located? 3) Can we identify in which gene the su mutation is

Genomic analyses reveal FoxG as an upstream regulator of wnt1 required for posterior identity specification in planarians

Embryonic specification of the first body axis requires the formation of an Organizer, a group of cells with the ability to instruct fates in the surrounding tissue. The existence of organizing regions in adults, i.e. during regeneration, which also requires patterning of new tissues, remains unstudied. To that aim, we study regeneration in planarians, flatworms that can regenerate any missing structure, even the head, in a few days. In planarians, as described in embryonic models, the cWNT pathway specifies the anterior-posterior axis. During the first 12-24h after amputation both wnt1 and notum (a Wnt inhibitor) are expressed in any wound, but 48 hours later they become restricted to posterior or anterior facing wounds, forming the anterior and the posterior organizers, respectively. In this study we undertook a genomic approach to further understand the mechanism that triggers the early expression of wnt1 and the specification of the posterior identity. Through ATAC-sequencing and CHIPmentation techniques we uncovered Cis-Regulatory Elements of Schmidtea mediterranea genome and analyzed them in notum and wnt1 (RNAi) animals. The result shows that already at 12 hours after amputation the chromatin structure of the wounds has changed its conformation according to the polarity of the pre-existing tissue. Analysing the DNA binding motives present in the proximal regulatory regions of genes down-regulated after wnt1 (RNAi) we found a few genes containing a TCF binding site, which include posterior Homeobox genes and chromatin remodelling proteins, suggesting that those are direct targets of the cWNT pathway and the responsible to trigger the expression of the posterior effectors. Furthermore, we have identified FoxG as an up-stream regulator of wnt1 transcription, probably though binding to an enhancer found in its first intron. Silencing of foxG inhibits the early phase of wnt1 expression and phenocopies the wnt1 (RNAi) phenotype, indicating its early role in specifying posterior versus anterior identity. Moreover, we have created a new open platform to interpret all transcriptomic and genomic results obtained (https://compgen.bio.ub.edu/PlanNET/planexp)