Discovery of four Noggin genes in lampreys suggests two rounds of ancient genome duplication

Abstract

The secreted protein Noggin1 was the first discovered natural embryonic inducer produced by cells of the Spemann organizer. Thereafter, it was shown that vertebrates have a whole family of Noggin genes with different expression patterns and functional properties. For example, Noggin1 and Noggin2 inhibit the activity of BMP, Nodal/Activin and Wnt-beta-catenin signalling, while Noggin4 cannot suppress BMP but specifically modulates Wnt signalling. In this work, we described and investigated phylogeny and expression patterns of four Noggin genes in lampreys, which represent the most basally divergent group of extant vertebrates, the cyclostomes, belonging to the superclass Agnatha. Assuming that lampreys have Noggin homologues in all representatives of another superclass of vertebrates, the Gnathostomata, we propose a model for Noggin family evolution in vertebrates. This model is in agreement with the hypotheses suggesting two rounds of genome duplication in the ancestor of vertebrates before the divergence of Agnatha and Gnathostomata.

Fig. 1. Phylogenetic analysis (a) and synteny (b) of lamprey Noggins.

A - Phylogenetic tree of Noggins is constructed using the Maximum Likehood algorithm. AC Anolis carolinensis, BF Branchiostoma floridae, CI Ciona intestinalis, DR Danio rerio, GG Gallus gallus, HS Homo sapiens, LF Lampetra fluviatilis, LC Lethenteron camtschaticum, PM Petromyzon marinus XL Xenopus laevis, XT Xenopus tropicalis. B - synteny of Noggin genes of sea lamprey P. marinus (PM) and western clawed frog X. tropicalis (XT).

Fig. 2. The expression dynamics of NogginA, NogginB, NogginC and NogginD in the early stages of development of L. fluviatilis.

Stage numbers are indicated according to ref. .

Fig. 3. Expression patterns of L. fluviatilis NogginA (a–j), NogginC (k–r) and NogginD (s–v).

a At stage 17 NogginA is expressed in the caudal chordamesoderm. b, c At stage 19 expression is in the entire chordamesoderm except for its most anterior part and in the dorsal region of the presumptive brain, excluding the most anterior part. d At stage 22 expression is in the floor and roof plate of the neural tube, in the notochord, in the somites, in the cheek process and in the diencephalon, midbrain and hindbrain. e–j At stages 23–25 expression pattern includes somites (i, j), upper and lower lips (g), notochord, in several areas of the brain: at the border of telencephalon and diencephalon, in zona limitans intrathalamica, at mid-hind brain boundary and in the hindbrain (g, i). k, l At stage 18 NogginC is expressed in the 2 chordamesoderm regions: anterior part and caudal part. m At stage 22 NogginC is found in the notochord, neural tube, pharyngeal arches, otic pits, cheek process and somites. n At stages 24 the expression of NogginC appears in the forebrain and otic visicles, upper and lower lips. o–q At stages 25 the cells of the pineal gland (transverse section on P) also express NogginC. r NogginC expression in somites at stage 25. s, t At stage 17 NogginD is expressed in the neural tube. u, v At stage 17 the expression of NogginD develops throughout the neural system in a diffuse manner and weakly in somites. as anterior intraencephalic sulcus, ch optic chiasma; di diencephalon, e eye, fb forebrain, h hindbrain, hb habenula, hpt hypothalamus, ll lower lip, ul upper lip, mes mesencephalon, MHB mid-hindbrain boundary, n notochord, ncc neural crest cells, np neural plate, nt neural tube, oc otic cup, op olfactory placode, pg pineal gland, pha pharyngeal arche, tel telencephalon, ZLI zona limitans intrathalamica.

Fig. 4. Matching of expression patterns of NogginA (a–j) and NogginC (k–t).

a At stage 25 NogginA is expressed in somites, especially in dorsal and ventral angles, notochord, hypochord, neural tube. b, c Transverse sections, the section levels are shown in (a). d Whole mount expression of NogginA at stage 27. e–g Saggital section of embryo of stage 27 reveals the expression of NogginA at the border of telencephalon and diencephalon along anterior intraencephalic sulcus, in zona limitans intrathalamica and at mid-hind brain boundary and in the eyes. h–j In the tailbud the expression of NogginA increases in the notochord of the growing tail. The levels of transverse sections are shown in (h). k–m At stage 25 NogginC is expressed in somites, notochord, neural tube and premigratory neural crest cells. l, m Transverse sections, the section levels are shown in (k). n Whole mount expression of NogginC at stage27. o–q Saggital sections reveal the stronger expression in the ventral part of the telencephalon and weak expression in ZLI and in otic cups. r–t at stage 27 NogginC is expressed in somites, notochord, neural tube at the trunk level (s), but in the growing tail (t) it reveals at a high level in the central (ventricular) zone of the neural tube. The levels of transverse sections are shown in (r). For abbreviations see Fig. 3.

Fig. 5. Expression pattern of L. fluviatilis NogginB.

a, e At stage 17 NogginB is expressed in the anterior end of the neural tube. b, f At stage 18 its expression is enhanced in the presumptive forebrain and hindbrain and weakens in the dorsal part of the presumptive midbrain. c, d, g, h At stage 19–21 the NogginB expression focuses on the presumptive forebrain (c, g) and in the most dorsal cells of neural tube (D, H) presumably - premigratory neural crest cells. i–r At stage 22–25 NogginB expression persists in the telencephalon and in ventral diencephalon and as is shown on transverse sections (q, r) in the premigratory neural crest cells. s Whole mount expression of NogginB at stage 27. t Saggital section of embyo of stage 27 reveal NogginB expression in the dorsal and ventral parts of the telencephalon and in the preoptic region of the hypothalamus. u–w horizontal section of embyo of stage 27. The section levels are shown in S. x, y NogginB expression at stage 28. For abbreviations see Fig. 3.

Fig. 6. Functional conservatism of Noggins in vertebrates.

Microinjections of mRNAs of lamprey NogginA (a, b), NogginB (c, d) and NogginC (e, f) in embryos of X. laevis causes the formation of secondary axes including anterior-headed structures like mRNA of X. laevis Noggin2 (i, j). For percentage of secondary axis and secondary head structures see Supplementary Figure 3. NogginD (g, h) does not induce secondary axes.

Fig. 7. Three possible scenarios of the evolution of Noggin genes in vertebrates.

a and b scenarios suggest one ancestral Noggin gene passed through two rounds of duplication (2 rounds of whole-genome duplication in a vs local+whole-genome duplications in b). c scenario suggests two ancestral Noggins passed through one round of whole-genome duplication.