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FGF signaling promotes precursor spreading for adult adipogenesis in Drosophila

Yuting Lei, Yuwei Huang, Ke Yang, Xueya Cao, Yuzhao Song, Enrique Martín-Blanco, José C. Pastor-Pareja

Preprint posted on 21 April 2022 https://www.biorxiv.org/content/10.1101/2022.04.21.489019v1.article-info

Out with the old, in with the new: during fly metamorphosis, the adult fat body develops de novo with a hand from FGF signalling.

Selected by Anastasia Moraiti

Background

Despite often being found in the same sentence as “excess”, and before “how to lose”, adipose tissue, or fat, plays essential roles in energy storage and release, highlighted by the development of insulin resistance or other metabolic complications in the absence of functional adipocytes. Despite its important roles, and our passion to eliminate it, we still have a poor understanding of the origins of fat.

The fruit fly Drosophila melanogaster is no exception in storing caloric excess in the form of adipose tissue, known as the fly fat body. The Drosophila fat body comes in two variants: larval and adult. The larval fat body descends from mesodermal precursors that are specified during embryogenesis and is eventually eliminated during metamorphosis, with only a few larval fat body cells persisting until the first few days after eclosion. However, the origins of the adult fat body are much less clear: does it come from the persisting larval cells or is there de novo adult adipogenesis? In this study, Lei et al., identify the adult fat body precursors during metamorphosis and characterise their expansion and differentiation. They also describe the essential role of FGF for this process.

 

Main findings

  • Using a GAL4 enhancer trap line identified as a tool to visualise the adult fat body in late metamorphosis (OK6-GAL4), the authors perform live imaging during early pupal stages and note the first OK6-positive cells arriving from the thorax at 15 h after puparium formation (APF). They denote these cells as the adult fat body precursors. These cells then expand towards the ventral abdominal epidermis (Figure 1), converge at the ventral midline at 30 h APF, and then spread laterally and dorsally towards the dorsal midline, forming at confluent monolayer at 65 h APF. During late metamorphosis, the authors observe the differentiation of the precursors into mature adipocytes, characterised by the accumulation of lipid droplets and by cell fusion that gives rise to the multinucleated adult adipocytes that have previously been described. To understand the origin of these precursors, they perform lineage tracing. Tracing with Mef2 in first instar (L1) larvae marked the adult adipocytes and adult muscles, but not the larval fat body. This indicates that the adult fat body diverges at L1 and shares a common progenitor with muscle cells.

Figure 1. Stills from a time lapse recording of an OK6-GAL4 pupa (ventral view), showing the migrating and proliferating population of the OK6-positive cells.

  • Lei et al. identify the GATA-like transcription factor Serpent (Srp), that is known to be necessary for larval fat body development, as essential for adult fat body as well. Using RNAi against srp they show that srpi flies develop a fat body, but a much reduced one, showing that srp is involved in the amplification rather than the specification of adult adipocytes.
  • The authors then turn to the role of FGF signalling in adult adipogenesis. They find expression of the FGF receptor heartless (htl) in the OK6-positive cells, and of the FGF ligand Thisbe (ths) in the underlying abdominal epidermis. Through loss of function of htl and ths, they demonstrate the requirement for FGF signalling, as loss of this signal resulted in a largely reduced fat body (although knockdown of pyr, another FGF ligand did not have an effect on the fat body). To understand the specific role of FGF signalling in this process they analyse fat body migration during Htl knockdown/overexpression in the OK6-positive cells. In both cases they observe reduced directionality in the migration of the fat body precursors, consistent with the previously described role of FGF as a chemoattractant cue (Sun et al., 2020). Furthermore though, they observe that precursors lacking functional htl often detach from the substrate leading eventually to their decreased numbers (Figure 2).

Figure 2. Left: angle of migration with respect to the AP axis (imaged 52-54 h APF) and Right: number of precursors (imaged 48-58 h APF) in wild-type and htl knockdown/overexpressing pupae. FGF signalling affects the directionality of migration and the attachment of the precursors to the substrate, as illustrated by the stationary number of precursors.

  • Finally, the authors probe the physiological roles of the adult fat body, by analysing flies where this tissue is reduced, after knockdown of srp or htl under OK6-GAL4. They highlight the role of the fat body as an essential energy reserve via their observation of ectopically accumulated lipid droplets in flies lacking a fat body, and via starvation assays. When comparing the survival of wild type and srp or htl flies after starvation, they observe that flies with a functional fat body displayed increased resistance to starvation.

 

Why I chose this preprint

This preprint is an important contribution to the field of fat body development by identifying the precursors of the mature adult adipocytes, and by showing that these come from a different lineage to the L1 larval fat body. Thus, it provides strong evidence for de novo adipogenesis rather than for derivation of the adult fat body from persisting larval cells. Another very interesting finding while investigating the role of FGF signalling during fat body expansion, is that the authors draw interesting parallels to mesoderm expansion during embryonic development. Here, as well as the role of FGF as a chemoattractant, they observe an important role in the attachment of the migrating precursors to the substrate. Finally, they are able to offer evidence for the functional importance of the fat body in the adult.

The system used in this study will be useful for further studies into fat body development. The Drosophila pupa is very amenable for live imaging, as the animal is sessile at this stage of development, and the fat body is very accessible as it lies right underneath the developing epidermis. The well-established array of genetic tools in Drosophila also comes in handy here, as shown by the very nice lineage tracing experiments, and the generation of viable flies lacking a fully developed fat body. These flies could also be used for further studies in fat body physiology.

 

Questions for the authors

  • You first identify the OK6-GAL4 cells at 15 h APF. Have you looked for this signal earlier during pupal stages or during late larval life?
  • Have you performed your RNAi experiments using a temporally controlled system (Gal80ts) to look for the specific effects of your knockdowns during larval vs pupal stages?
  • How can your system be used for further studies on alternative roles of the adult fat body, for example in systemic immunity or regulation of reproduction?

 

 

Tags: adipogenesis, drosophila, fat body

Posted on: 10 May 2022

doi: https://doi.org/10.1242/prelights.31958

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Author's response

José C. Pastor-Pareja shared

José Pastor-Pareja shared

1. You first identify the OK6-GAL4 cells at 15 h APF. Have you looked for this signal earlier during pupal stages or during late larval life?

We were very lucky to discover that OK6-GAL4 labeled adult fat body precursors. The question of the origin of the adult fat body is a decades-old mystery that Alberto Ferrus and Peter Lawrence already tried to solve back in the 80’s. Our study reveals important details about the development of the adult fat body, crucially a lack of relation with the larval fat body. However, part of the mystery still remains because, answering your question: yes, we have looked for those cells earlier than 15 h APF, but we can’t find them. It seems that OK6-GAL4 turns on at about that time, perhaps a little earlier, at 12 h APF. Therefore, we don’t know the exact location of these cells in the late larva or the early pupa. Our guess, same as Deborah Keiko-Hoshizaki’s in the 90’s (but for slightly different reasons), is that the place to look for these cells would be the myoblast-rich adepithelial populations that attach to the wing and leg imaginal discs, but so far we haven’t been able to prove it.

2. Have you performed your RNAi experiments using a temporally controlled system (Gal80ts) to look for the specific effects of your knockdowns during larval vs pupal stages?

Yes, we performed a few experiments like this with GAL80ts and we don’t see any effect when knock down is restricted to the larval period. This fits well with the fact that OK6-GAL4 may not be expressed in precursors earlier that 12-15 h APF, as mentioned in the previous answer. Results of other experiments with GAL80ts show that FGF signaling is continuously required during the migration period in mid and late pupal stages.

3. How can your system be used for further studies on alternative roles of the adult fat body, for example in systemic immunity or regulation of reproduction? 

Knowing how the adult fat body is assembled during metamorphosis opens the door to specifically manipulate it and dissect its roles from those of the larval fat body. Importantly, we can now perform the most basic experiment of all: ablate and see what happens. We did a little bit of that already in this study and concluded that the adult fat body is an energy store that provides resistance to starvation. Proof of this did not exist until now, but I would not say this is a revolutionary finding, as a storage role is highly expected from an adipose tissue. However, for both insects and mammals, the adipose tissue is increasingly seen less as a simple energy store and more as a complex endocrine organ with multiple regulatory roles in almost all aspects of physiology. A good starting point to uncover or dissect those roles is to study flies without fat body. Flies without fat body, in addition, may be a good way to model lipodystrophy, a word that designates a large number of disease conditions in which the adipose tissue degenerates or does not develop.

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