A nemertean excitatory peptide/CCHamide regulates ciliary swimming in the larvae of Lineus longissimus | Frontiers in Zoology


Neuropeptides are neurotransmitters that are involved in the regulation of most behavioral and physiological processes in animals. Many neuropeptide systems are ancestral to bilaterians and orthologous neuropeptides are deployed in the different bilaterian lineages independent of their nervous system organization [13]. Only few of the orthologous neuropeptides are well conserved in their amino acid sequences between the different bilaterian lineages and it is often the orthology of their receptors, that reveals their homology [14]. For example, the protostome orthologs of vertebrate neuromedin -B/bombesin- and endothelin-related neuropeptides, the CCHamide and excitatory peptide (EP), are neuropeptides where the orthology of the deuterostome and protostome peptidergic systems could only be detected because of their receptor similarity [2, 3, 5].

The EP was initially discovered in the earthworms

Eisenia foetida

and

Pheretima vittata

[

6

] and has since been identified in many annelid and mollusk species. It has been described under various names depending on the taxon and due to either its myo-excitatory effect or its C-terminal structure (see Table 

1

for species, peptide names and references). The arthropod ortholog CCHamide was first discovered in the silk worm

Bombyx mori

[

23

] and is known from various arthropods, including insects [

3

,

23

,

24

], crustaceans [

3

,

24

26

], myriapods [

27

] and chelicerates [

24

,

28

,

29

], with its nomenclature based on the presence of two conserved cysteine residues and an amidated C-terminal histidine residue. Because of the presence of the two cysteine residues, the usually amidated C-terminus and a similar precursor structure in CCHamides and EPs (Fig. 

1

a), the two peptides were already recognized as possible orthologs [

2

,

3

,

17

] before the corresponding receptors were known. This orthology hypothesis was then confirmed with the deorphanization of their orthologous receptors [

12

,

24

]. Notably, the CCHamide system duplicated within the insect lineage into two distinctive CCHamides with two distinctive CCHamide receptors, where each of the two peptides seems to specifically activate its own, corresponding receptor paralog [

5

,

24

]. Experiments showed that CCHamide is involved in the regulation of feeding, sensory perception and the control of insulin like peptides in

Drosophila melanogaster

[

36

38

] and connected to feeding in other insects [

5

,

39

]. Its expression in different larval or adult insects is connected to the digestive system [

23

,

38

42

]. Experiments with EP showed a myo-excitatory effect that included digestive tissues of oligochaetes [

6

], leeches [

7

,

8

], and a gastropod species [

15

], and association with digestive tissues was also shown by immunohistochemistry on a polychaete [

10

]. (See also Table 

2

for functional and anatomical association of EP/CCHamides). The myo-excitatory effect and the expression of EP was observed on tissues of adult animals.

Table 1

Discovery and nomenclature of trochozoan EPs

Annelida

Eisenia foetida, Pheretima vittata

GGNG peptide

[6]

Hirudo nipponia, Whitmania pigra

GGNG peptide, LEP (leech excitatory peptide), EEP (earthworm excitatory peptide)

[7, 8]

Eisenia foetida

GGNG peptide, LEP, EEP,

[9]

Perinereis vancaurica

GGNG peptide, PEP (polychaete excitatory peptide)

[10]

Capitella teleta

(Hirudo japonica, Hirudo medicinalis, Alvinella pompejana)

GGNG

[11]

EP (excitatory peptide)

[2]

Platynereis dumerilii

EP (excitatory peptide)

[12, 13]

Dinophilus gyrociliatus, D. taeniatus, Trilobodrilus axi

EP

[14]

Mollusca

Thais clavigera

GGNG peptide, TEP (Thais excitatory peptide)

[15, 16]

Lottia gigantea

GGNG

[17]

Crassostrea gigas

GGNamide

[18]

Theba pisana

GGNG

[19]

Sepia officinalis

GGNG, GGNamide

[20]

Charonia tritonis

GGNG

[21]

Patinopecten yessoensis

GGNamide

[22]

Fig. 1

Protostome EP/CCHamide sequences. a Schematic structure of the 116 amino acid long L. longissimus EP2 precursor. Scale bar on the upper right indicates the length of 10 amino acids. b Alignment of the predicted EP/CCHamide peptides of different protostomes with the phylogenetic relationship of the different taxa. C-terminal GKR, GKK or GRK residues indicate the precursor cleavage site and a C-terminal amidation of the residue N-terminal to the glycine, a missing glycine residue (e.g. M. tardigradum) indicates only cleavage without amidation. Peptide sequence logo was created from the alignment. Phylogeny is depicted according to Dunn et al. 2014 [30], annelid phylogeny according to Struck et al. 2015 [31] with Orthonectida as an annelid taxon [32], arthropod phylogeny according to Yeates et al. 2016 [33] and nemertean phylogeny according to Andrade et al. 2014 [34] and Kvist et al. 2015 [35]. Dashed line indicates unclear relationship, asterisks indicate the heteronemertean branch. Full precursor sequences are available in Additional file 1

Table 2

Association of EP/CCHamide peptidergic signaling based on expression, peptide detection and functional analysis of previous studies

Annelida

Eisenia foetida, Pheretima vittata

Isolation of EP from gut tissue as well as whole bodies and excitation of gut tissue by EP application.

[6]

Hirudo nipponia,

Excitation of the crop gizzard by EP application.

[7]

Eisenia foetida

EP binding capacity is high in anterior part of digestive tract and the nephridia.

[9]

Whitmania pigra

EP immunoreactivity in supra-esophageal ganglion, circum-esophageal connective, sex segmental ganglion.

[8]

Perinereis vancaurica

EP immunoreactivity in CNS, epithelial cells of pharynx and epidermal cells.

[10]

Mollusca

Thais clavigera

Excitation of esophagus and penial complex by EP application, EP immunoreactivity in CNS and nerve endings of the penial complex.

[15]

Thais clavigera

EP1 expression in sub-esophageal, pleural, pedal and visceral ganglion and EP2 expression in pedal and visceral ganglion.

[16]

Hexapoda

Bombyx mori – larvae

CCHa expression in the central nervous system and the midgut.

[23]

Phormia regina – adults

CCHa2 injection stimulates feeding motivation (measured by the proboscis extension reflex at different sugar concentrations).

[5]

Delia radicum – larvae

CCHa1 was exclusively detected in the gut.

[41]

Spodoptera exigua – larvae

CCHa1 and CCHa2 are expressed in the larval gut and brain. Starvation increased CCHa1 expression in larvae.

[39]

Drosohila melanogaster – larvae & adults

High CCHa2 expression in gut and low expression in brain; high CCHa2 receptor expression in brain and low expression in gut.

[40]

D. melanogaster – adults

Upregulation of CCHa (1?) in the brain of starved animals. RNAi knockdown of the CCHa1 receptor and CCHa1 receptor mutants showed an abolishment of a starvation-induced increase in olfactory responsiveness.

[36]

D. melanogaster – larvae & adults

Distinct CCHa1 and CCHa2 immunoreactivity in the digestive tract in both larvae and adults.

[42]

D. melanogaster – larvae

CCHa2 is highly expressed in fat body and slightly in gut, CCHa2 receptor is expressed in few endocrine cells in the brain including insulin like peptide (ILP) 2 producing cells. Starvation reduces CCHa2 expression. CCHa2 receptor mutants showed no change in ILP 2 and 3 expression but reduced ILP 5 expression. CCHa2 mutants show growth retardation and developmental delay. CCHa2 mutants show reduced ILP 5 expression and reduced body weight.

[38]

D. melanogaster – larvae & adults

Larvae: CCHa2 mutants show reduced feeding rate/activity and have a delayed development. Larvae and Pupae have reduced expression of insulin like peptide 2 and 3. CCHa2 is highly expressed in gut and slightly in brain. No effect detected for CCHa1.

Adults: CCHa2 mutants show reduced feeding and reduced locomotory activity. No effect detected for CCHa1.

[37]

Crustacea

Marsupenaeus japonicus – juvenile/adults

Highest expression of CCHa in ventral nerve cord, brain, eyestalks and gills, only low expression in intestines and stomach tissue. No effect of starvation on CCHa expression.

[43]

Nephrops norvegicus – adults

Tissue specific transcriptome detection of two CCHa’s in brain, thoracic ganglia and eyestalks, but not in hepatopancreas or ovaries.

[26]

Annelids and molluscs, as well as other closely related taxa such as brachiopods and nemerteans, also possess planktonic larvae that usually metamorphose into morphologically different, benthic adults. This ciliated larva is the name giving characteristic of the spiralian taxon Trochozoa [30, 4447]. While antibodies against neuropeptides (usually FMRFamide) have been used widely to describe the nervous system of trochozoan larvae [4856], there are comparably few studies that investigated the behavioral effect of neuropeptides in such larvae [5761] and neither behavioral nor immunohistochemical studies investigated the EP/CCHamide in trochozoan larvae. In nemerteans no functional studies with neuropeptides have been reported so far. One group of nemerteans, the pilidiophorans [34, 35, 62], have a planktotrophic and long living pilidium larva (with a few exceptions [63, 64]). Within such a pilidium larva, the juvenile worm develops from initially isolated imaginal discs that eventually fuse and overgrow the larval gut until it hatches after several weeks and often devours its own larval tissue [64, 65]. The larval nervous system lacks a typical neuropil and comprises an apical organ that consists of an apical plate with densely arranged secretory cells at the base of a prominent apical tuft, and neurons that are associated with the digestive system and a nerve net that loosely covers the body and eventually innervate a prominent nerve underneath the ciliary band [51, 65, 66].

Here we report on the evolution of EP/CCHamide orthologs in ecdysozoans and spiralians with a focus on nemerteans. We test the activation of a single EP receptor by two EP splice variants in the nemertean Lineus longissimus, the EP expression in the pilidium larvae and juveniles of L. longissimus, and the influence of the EP on the behavior of L. longissimus larvae.



Source link

Notice: compact(): Undefined variable: limits in /customers/6/d/3/sciencetells.co.uk/httpd.www/wp-includes/class-wp-comment-query.php on line 853 Notice: compact(): Undefined variable: groupby in /customers/6/d/3/sciencetells.co.uk/httpd.www/wp-includes/class-wp-comment-query.php on line 853