This book systematically introduces the basic knowledge of the Hypothalamic Pituitary Gonads (HPG) axis and provides information on the location and regulation, gene mutations, function, reproduction, life cycle, sexual behaviour, disorders and the role of the environmental factors on GnRH gene. Also, we focused on gene and receptor structures, and the signalling pathways of GnRH, and its related genes and hormones such as Luteinizing Hormone (LH), Follicle-Stimulating Hormone (FSH), Progesterone (P4) and Oestradiol (E2). Also, it was pointed to Gonadotropin Inhibiting Hormone (GnIH) and its related peptides, such as RFamide peptides which were found to decrease hormones secretion by working on HPG in an inhibiting biosynthesis process of gonadotropin (α–β) subunits. In addition, the roles of hormones on fertility and reproduction, also, disruption resulted from mutations. Special characteristics of many hormones and pulsatile secretions of GnRH were summarized.
This information is necessary for an understanding way of work some necessary hormones also, elucidating reasons infertility. Although, several studies have presented a lot of kinds of regulators of expression for many genes related by fertility in many species, however, the knowledge of the relationship between molecular relating and infertility still has many gaps and problematic, also, many remains to be discovered, and a lot of significant questions (such as sexual behaviour, the regulation of gonadotropin secretion, and growth of tumour cell, in addition to; whether GnRHs are expressed as a full-length or not?), need to be solved, and mechanisms of pulsatile secretions and potential functions of many reproductive hormones need to be elucidated. So, this investigation very important for those researchers and readers to start solve and answer these questions and understanding many things related to (HPG) axis, and (GnRH).
LIST OF CONTENT
ABSTRACT
1. INTRODUCTION
2. GONADOTROPIN-RELEASING HORMONE (GnRH)
2.1. Gonadotropin-releasing hormone (GnRH) signalling pathway
2.2. Gonadotropin-Releasing Hormone gene (GnRH gene)
2.3. Gonadotropin-Releasing Hormone Receptor (GnRH-R)
2.4. GnRH peptides and receptors
2.5. GnRH gene Transcription Regulation
2.5.1. Oct-1, Oct-2, Pit-1and Unc86
2.5.2. Dlx-2 and Msx-1
2.5.3. Pbx-1
2.5.4. Homeobox protein (Otx-2)
2.5.5. GATA-A and GATA-B
2.5.6. PKC, Jun, Fos, AP-1, and NK-B
2.5.7. Pur-A, Pur-B, Brn-2, hnRNP-A/B, and hnRNP-G
2.5.8. Kisspeptin
2.6. Expression of the GnRH-R gene
2.7. Effects of GnRH activation of GnRH-R and its signalling pathways
3. GONADOTROPINS (Gn)
3.1. Luteinizing hormone (LH)
3.1.1. Regulation of LH (LH-β) by GnRH
3.1.2. Regulation of LH-β by Follistatin (Fo), Activin (Ac), and Inhibin (In)
3.1.3. Regulation of LH-β by Estrogen (E) and Androgen (An)
3.2. Follicle-Stimulating Hormone (FSH)
3.2.1. Regulation of FSH (FSH-β) by GnRH
3.2.2. Transcriptional regulation of FSH-β encoding genes
4. PROGESTERONE (P4) AND OESTRADIOL (E2)
5. SUMMARY POINTS LIST
6. CHALLENGES AND FUTURE DIRECTIONS
7. LITERATURE CITED
7.1. RELATED RESOURCES
Abbildung in dieser Leseprobe nicht enthalten
ABSTRACT
This book systematically introduces the basic knowledge of the Hypothalamic Pituitary Gonads (HPG) axis and provides information on the location and regulation, gene mutations, function, reproduction, life cycle, sexual behaviour, disorders and the role of the environmental factors on GnRH gene. Also, we focused on gene and receptor structures, and the signalling pathways of GnRH, and its related genes and hormones such as Luteinizing Hormone (LH), Follicle-Stimulating Hormone (FSH), Progesterone (P4) and Oestradiol (E2). Also, it was pointed to Gonadotropin Inhibiting Hormone (GnIH) and its related peptides, such as RFamide peptides which were found to decrease hormones secretion by working on HPG in an inhibiting biosynthesis process of gonadotropin (α–β) subunits. In addition, the roles of hormones on fertility and reproduction, also, disruption resulted from mutations. Special characteristics of many hormones and pulsatile secretions of GnRH were summarized.
Keywords; HPG, GnRH, GnRHR, and Gonadotropins.
1. INTRODUCTION
The Hypothalamic Pituitary Gonadal axis (HPG) plays important roles in steroids production, body development, reproductive activities and ageing in male and female. They also participate in the regulation of a number of the body systems, such as the immune and reproductive systems (66). Therefore, controlling the release of some important hormones like; Gonadotropin-Releasing Hormone (GnRH), Luteinizing Hormone (LH), Follicle-Stimulating Hormone (FSH), testosterone and Estrogen (E) either directly or indirectly should be beneficial to several biological body activities (136; 199). GnRH is secreted from the hypothalamus through GnRH-expressing neurons. Thus, the anterior portion of the pituitary gland produces two basic hormones LH and FSH, then successively the gonads are stimulated to produce sex hormones, testosterone in males and estrogen in females (15).
The HPG axis consists of 3 levels: hypothalamus, pituitary, and gonads (ovaries in females and testes in the males). GnRH, LH and FSH together with sex hormones are all products of HPG axis. Moreover, differentiation and maturation of reproductive organs, steroids excretions, spermatogenesis in mature males and oogenesis in mature females are all mediated via several reproductive hormones controlled by HPG, Figure (1) (66).
However, a few years ago, a family of neuropeptides has been discovered and was recognized as the RFamide-related peptides (RFRPs). They are characterized by the site [Arg-Phe-NH2] motif at C-terminal extremities (5). Two groups of RFRPs proved to participate in the activities of the HPG axis: Gonadotropin Inhibiting Hormone (GnIH) and its related peptides. These RFRPs were found to decrease hormonal secretion by inhibiting HPG biosynthesis of gonadotropin (Alpha–Beta) subunits in a dose-dependent manner (179; 186). Also, two RFRPs; RFamide-related peptide-one, RFRP-1 and RFamide-related peptide-three, RFRP-3 were isolated from the brain in human (185), bovine (185), rhesus macaque (185) and rats (67), whereas RFRP-3 was found to inhibit LH release in cattle (70), sheep (25), and rats (67). Neurons expressing RFRP-3 have located in the dorsomedial region of the human brain and their cells become in contact with GnRH neurons in the preoptic position (184). Also, Tsutsui et al. (179) discovered a new RFamide-related peptide in the hypothalamus of Japanese quail. Finally, those RFRPs were designated as Gonadotropin Inhibiting Hormone (GnIH). Also, GnIH and other peptides have been detected in the brain of fish, amphibians, and avian species (6; 47; 156; 184; 188).
The objectives of this book are to present a comprehensive summary of the regulatory actions of GnRH as a gene, and as a hormone along with the specific endocrine receptors in human and animals including cattle, sheep, and pig. The comprehensive and basal information declared in this article may supply a better understanding of the function of the Hypothalamic Pituitary Gonadal (HPG) axis which is principaly useful for many researchers and students. We were so intent to make comparisons with other reviews which focused mainly on HPG axis, using one or two hormones or probably on all hormones but in one sex only. The current book focused on all hormones included HPG axis in both male and female. The genetic pathway and the structure of genes related to the HPG axis were also under discussion.
Abbildung in dieser Leseprobe nicht enthalten
Figure 1
Levels of The Hypothalamic Pituitary Gonadal axis (HPG): hypothalamus, pituitary, and gonads (ovaries in the female and testes in the male).
2. Gonadotropin-Releasing Hormone (GnRH)
Gonadotropin Releasing Hormone, the some called it; GnRH, LHRH. It made by (Hypothalamus). GnRH important to make and release the (LH) and (FSH). In the male, (LH) and (FSH) stimulate the testicles to make testosterone. In the female, those hormones stimulate the ovaries to make females` hormones like; (P4) and (E) (5; 15; 66).
2.1. Gonadotropin-releasing hormone (GnRH) signalling pathway
Widmann et al reported that; GnRH signalling is related by the divergent growth at puberty (especially at the onset) under the influence of two important connected hubs; Diacylglycerol-Kinase-eta gene (DGKH) and Betacellulin gene (BTC). Both genes react directly with the GnRH signalling pathway (202). Moreover, the gene interaction network non-SMC condensin I complex-subunit-G (NCAPG) included fourteen connected genes, organized in five groups descended according to their connectivity group-1 including; MRVI-1, PTPRD, NCAPG, FSHR, BRE and ACCN1 genes (59) group-2 including; ALK gene (59) group-3 including; SMG-6, PBX-4, GRIK-2 and ASB-5 genes group-4 including; AGTPBP-1 gene (135) and group-5 including; POLR-2G and ADAM-15 genes (26). such discovered novel results concerning the functional impact of NCAPG on growth (202).
BTC gene belongs to Epidermal Growth Factor family (EGF) which participate in differentiation and proliferation of cells and therefore stimulates growth (39). It binds to EGF to form the EGF receptor (EGF-R). Moreover, growth promoting impact on mesenchymal cells (40) and its effect on growth are mediated through interaction with EGF-R (202). Also, Watanabe et al. (198) proved that the latter receptor is a primary receptor for BTC. Studies on ovarian follicles indicated that there is an interaction between LH and BTC during maturation of follicles. Which allow BTC to be a downstream mediator to propagates LH signals (29; 141). The gene DGKH which belongs to Dlacylgly cerol kinases enzymes family (115) was found to affect the divergent growth (202) and proved to be a second candidate for regulation of the Raf/Ras/ERK/MEK signalling flow that affected the growth regulation processes including differentiation, proliferation, and transformation (202). The role of C-Raf in growth and development appeased in mutant rats. The necessary roles of (C-Raf) for growing and development appear in (C-Raf) mutant in rats (204), Figure (2).
BTC and DGKH are two out of the ten most densely connected genes within the interaction network. The other genes are TH1-like (TH1L) Drosophila, Murine retrovirus integration site-1 (MRVI-1), Ubiquitin specific peptidase-40 (USP40), Fragile histidine triad (FHIT), DnaJ (Hsp40) homolog subfamily B, member-14 (DNAJB-14), Brain and reproductive organ-expressed (BRE), Syntaxin-12 (STX-12), and Prolactin-releasing hormone receptor (PRLH-R) (9; 202; 215). Both BTC and DGKH are encoding proteins that established a binding process to components in the GnRH signalling pathway and they, also, have nodes highly connected to the network of growth. DGKH may interact with growth processes through the functional role of (C-Raf) (202), Figure (2).
Abbildung in dieser Leseprobe nicht enthalten
Figure 2
Gonadotropin-releasing-hormone signalling pathway including genes that are in the Partial Correlation Information Theory (PCIT), their effects and pathways.
2.2. Gonadotropin-Releasing Hormone gene (GnRH gene)
The identity of the GnRH gene was clarified (114). Eraly et al reported that twenty-four decapeptide GnRHs have been found in nerve tissues, sharing a highly conserved structure ( pGlu-His-Trp-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-NH2 (42). Four GnRHs and one GnRH receptor (GnRH-R) are expressed in several mammals, whereas thirteen GnRHs and many GnRH-Rs have been discovered in several non-mammalian vertebrates (42; 131) The sequencing of the nucleotide for human genome provides a chance to identify which of these GnRHs and their receptors may be expressed in primates (131). Out of four GnRHs expressed in primates, only GnRH-I was expressed in mammals and GnRH-II in chicken but all were discovered in the genome of humans (78).
GnRH-I gene is located on chromosome 8 in mammals and in humans it is located on chromosome 8 p11.2–p21. GnRH-I gene encodes a preproprotein that is proteolytically processed to generate a peptide that is a member of the GnRH family of peptides. GnRH-II gene is located on chromosome 20 p13 (Start 3,043,622 bp to 3,045,747 bp) in humans (131).
The human GnRH-I gene consists of 4 exons separated by three introns and is present as a single gene copy on chromosome 8p11.2-p21 Figure (3A) (17; 144; 210). Exon number one is untranslated and consists of sixty-one base pair in messenger RNA (mRNA) in the hypothalamus. Exon number two; encodes the signal sequence, the GKR processing signal, the GnRH decapeptide, and the first eleven GAP residues. The third exon codes for the next thirty-two GAP residues. The fourth encodes the remaining GAP residues and contains the translation termination codon as well as 3`-UTR area. The human GnRH-II gene is located on chromosome 20p13. It is similar to the GnRH-I gene in the organization, the number of exons and numbers introns, however, the human GnRH-I gene (5 kb) is taller than GnRH-II (2.1 kb) Figure (3B) (1; 17; 144; 210). Moreover, GnRH-II gene is located on chromosome 17 in pigs Figure (3C).
The GnRH-1 gene is expressed in the hypothalamus (42), in discrete neurons (198) Also, the linear decapeptide end-product is synthesized from a 92-amino acids preprohormone in the preoptic anterior hypothalamus. Gonadoliberin-1 stimulates the release of (LH) and (FSH) hormones, which are important for reproduction. Mutations in this gene are associated with hypogonadism; thus GnRHs and their receptors are necessary for reproduction (48). GnRH-I gene was thought to be the only form of GnRH until 1984, however, after a few years GnRH-II was detected in the chicken brain (119) and GnRH-III was found in the lamprey brain (166).
The three forms of GnRH, differ in the sequence of amino acids, embryonic origin, and localization. It is interesting to note that GnRH-II and GnRH-I have similarly conserved amino acids, but are different from GnRH-III (66; 76). The cDNA for GnRH-II differs from that for GnRH-I by having Histidine, Tryptophan and Tyrosine residues in chicken (76; 119; 189), and is, also, conserved from primitive fish to humans. Also, GnRH-II serves as a potent inhibitor of K channels in the amphibian sympathetic ganglion (28; 116).
GnRH-II gene produces Progonadoliberin-II protein in humans. It is cleaved to secrete a ten- amino acid peptide hormone. This hormone regulates reproduction in females by stimulating the secretion of (LH) and (FSH) (8; 16; 100).
GnRHs are involved in several physiological processes in the body. For example, they influence the reproductive behaviour through neuromodulatory pathways and secretion of sex-hormones, also; GnRH-I controls the maturation of ovaries and testes through of gonadotropin release by the pituitary gland. Moreover GnRH-II and GnRH-III affect reproductive behaviour and related activities by neuromodulatory control of in the central nervous system (CNS) (57; 64), Figure(4).
Silveira et al. (162) reported that mutations in Fibroblast Growth-Factor R-1 (FGFR-1), Fibroblast Growth-Factor - 8(FGF-8), Kallmann syndrome (KAL-1), Prokineticin-2 (PROK-2), and Prokineticin Receptor-2(PROKR-2) are associated with the disruption of the migration and development of neurons for GnRH. This case leads to the Kallmann syndrome (Ks), which is known as the complicated genetic condition characterized by olfactory abnormalities and Idiopathic hypogonadotropic hypogonadism (IHH). Side by side; mutations in GnRH-I, GnRH-R, and KISS-1R genes lead to Normosmic-Idiopathic hypogonadotropic-hypogonadism (N-IHH) which interact with the action, secretion, and normal synthesis of GnRH (177).
Abbildung in dieser Leseprobe nicht enthalten
Figure 3
Two forms of human GnRH termed (A) - GnRH-I and (B) - GnRH-II, encoded by separate genes on chromosome 8p11.2-p21 and 20p13, respectively, cDNA and genomic structures for them. (C) - Exon/intron organization of porcine GnRHR-II genes and its derived amino acid sequences.
Abbildung in dieser Leseprobe nicht enthalten
Figure 4
Three types of GnRHs are helping and involving in physiological processes through many hormonal, and different
2.3. Gonadotropin-Releasing Hormone Receptor (GnRH-R)
The role of GnRH is mediated by GnRH receptors (GnRH-Rs), which belong to the G-protein coupled with rhodopsin-like receptor superfamily (GPCR) (17). Regulatory effect of GnRH-Rs is necessary because of the quantity of LH released in response to the physiological stress by GnRH is dependent on the concentration of GnRH-Rs on the plasma membrane of gonadotropes (203).
Stimulation of the pituitary gland by GnRH is required to maintain a steady-state concentration of GnRH-Rs and mRNA. In sheep; GnRH is released in a pulsatile fashion from the hypothalamus and is a homologous regulator for its own receptor (23; 24; 61). Unlike the effects of pulsatile GnRH, continuous infusion of it leads to a desensitization of gonadotropes. marked by about 50% decrease in the number of GnRH-Rs (120). It is not known whether the reduction in numbers of GnRH-Rs is caused by mediation at the level of expression of GnRH-R gene or receptor internalization, or both (30; 97; 183).
Clark and Mellon (22) reported that there are 3 GnRH-Rs, the first one was the GnRH-I receptor localized on chromosome 4, the second was a putative GnRH-II receptor gene found on chromosome (1), and the third was a sterile GnRH-II receptor-like homolog gene previously identified as being 40% identical to the GnRH-I receptor gene.
The GnRH-R is a member of GPCR superfamily or G-protein coupled receptor which includes a characteristic 7TM-R structure, unlike other many genes of G-protein-coupled receptors (GPCRs) the human GnRH-R gene consists of 3 exons separated by 2 introns and spans more than fifteen-kilo-pair along the chromosome (17; 31; 167). The GnRH-RI gene of human discovered by utilization of genomic southern blot and chromosomal in situ hybridization, also its found as a single copy on the chromosome (4) (43; 74). Exon number one of; the GnRH-RI gene contains the 5`UTR area and the first (five hundred twenty-one base pair) of the open reading frame, which encodes the first (3) TM domains and a portion of the 4th TM domain (2; 147). Exon number two; encodes the next (two hundred twenty base pair) of the reading frame, which encompasses the remainder of the 4th TM domain, the 5th TM domain, also, part of the 3rd intracellular loop. The third one consist of the rest of the coding sequence and the 3`UTR area Figure (5A). The human GnRH-RII gene at the chromosome (1). GnRH-RII gene similar to GnRH-RI gene in the number of exons and introns also, the same organization Figure (5B). The location of all the exons and introns boundaries of the GnRH-R gene in human is perfectly conserved in the ovine and rodent sequences, however, the first intron of the human gene is small (2; 74; 96; 147).
GnRH-RI lacking a carboxyl-terminal tail has been found in the human body Figure (6A) (17). GnRH-RII gene from a monkey that was (ninety-six percent) identical with the GnRH-RII gene in human. The GnRH-RII gene in monkey encoded a 379-amino acid G protein-coupled/seven transmembrane domain receptor (7TM-R) having a C-terminal cytoplasmic tail Figure (6B) (131).
In the human GnRH-R, many of amino acid remains of critical necessary for function have been identified. For example, but not limited, (Lys) [191] remain has been found to be an important determinant of the GnRH-R`s expression (46). Also, (Asp) [98], (Trp) [101], (Asn) [102], (Lys) [121], (Asn) [212], and (Asp) [302] are very necessary for ligand binding (46; 214). The (Ala) [261] in the 3rd intracellular loop is necessary for G-protein coupling and internalization for the receptor (3; 214). Furthermore, in several species of mammalian such as humans, sheep, cow, and chimpanzees, the GnRH-RII gene has been out of order through a mutation because of (frame-shift) led to; stop codon, thus, the GnRH-RII in these species has not expressed (117; 118; 131).
Abbildung in dieser Leseprobe nicht enthalten
Figure 5
Exon/intron organization of the human GnRHR-I and II genes and its derived amino acid sequences organized as the seven-transmembrane domain receptor (7TM-R), cDNA, and genomic structures for them.
Abbildung in dieser Leseprobe nicht enthalten
Figure 6
Comparison of the structures, 7TM of human GnRH-II receptor (A) and monkey GnRH-II receptor (B).
In pig the full-length for predicted 5-transmembrane (TM) GnRH-RII is two hundred eighty-six amino acids (AA) (NCBI accession number AAS68623.1) (105; 153). While, the full-length for GnRH-RII is three hundred seventy-seven (AA) (NCBI accession number AAS68622.1) and its forty-two percent homology to GnRH-RI, which is three hundred twenty-eight (AA) in length (NCBI accession number NP_999438.1) (37). Table (1), Figure (7). Also, the GnRH-RI and GnRH-RII are a member of the (Rhodopsin-like-receptors superfamily) of (GPCRs) containing the N-terminus add to 7-TM alpha-helical domains (173). There is a comparison between predicted 5-transmembrane (TM) GnRH-RII Figure (7A), GnRH-RI Figure (7B) and GnRH-RII Figure (7C) in the pig. The GnRH-RI and GnRHR-II connected via 3 extracellular loops (ECLs) and three intracellular loops (ICLs) Table (2), [Figures (7B), (7C)]. The remarkable thing here in a C-terminal tail`s GnRH-RII contains a fifty-two (AA), unlike, GnRH-RI, but the same case in non-mammalian GnRH-Rs [Figures (7B), (7C)] (37; 173).
The cytoplasmic tails are marketable among (GPCRs), also, promote rapid receptor desensitization (PRRD) and internalization (PRRI) (63; 111). It is worth mentioning, the GnRH-RII in monkey is fully desensitized to GnRH-II treatment after (1h) while, the tail`s GnRH-RI for human failed the desensitize to its receptor (GnRH-RI) agonist Triptorelin [(Triptorelin) is a gonadotropin-releasing hormone, also work as a potent inhibitor of the synthesis of estrogen (in females) and testosterone (in males)], through the period of entire sampling (1.5 h) (27; 37; 131). Thus, there are internalization differences between GnRH-RI and GnRH-RII in a cytoplasmic tail, where the GnRH-RII can use (Beta-arrestin) for internalization unlike GnRH-RI, that internalized without (Beta-arrestin) (105; 112; 153). In the C-terminus, the phosphorylation of serine residues (338) also (339) through G-protein-coupled receptors-kinases (GPCR kinases) is a necessary for (Beta-arrestin) independent internalization of GnRH-RII, while many other regions in the intracellular Loop-3 (ICL-3) and C-terminal tail are enough for (Beta-arrestin) dependent internalization [73, 74]. Thence, there are two distinct pathways for coordinate internalization of GnRH-RII [74]. Also, there are other remarkable differences between the GnRH-RI and GnRH-RII in the structure of residues that are conserved amongst the (GPCRs), where GnRH-RI has a distinct (Asn/Asp) domain, unlike the GnRH-RII that has an (Asp/Asp) small domain in transmembrane-2 and 7 (TM-2 and TM-7) (37; 105), Table (1).
Abbildung in dieser Leseprobe nicht enthalten
Figure 7
Structural comparison of GnRHRs; (A) - predicted 5-transmembrane (TM), (B) - GnRHR-I and (C) - GnRHR-II in the pig. GnRHR-I and RII are both 7-TM G-protein coupled receptors; however, GnRHR-I lacks a C-terminal tail present in GnRH-RII. The 5-TM GnRH-RII isoform lacks transmembrane (TM) domains (1) and (2) due to alternative splicing in exon (one) of the GnRH-RII gene, resulting in the coupling of a truncated extracellular N-terminus directly to transmembrane-3 (TM3). (Y) Is mean Glycosylation sites, while the darkened residues isoforms represent predicted sites of phosphorylation.
Abbildung in dieser Leseprobe nicht enthalten
Continue Figure 7
Structural comparison of GnRH-Rs in the pig.
[...]
- Arbeit zitieren
- Ahmed Saleh (Autor:in), Amr Rashadb (Autor:in), Nada Hassaninea (Autor:in), Yongju Zhaoa (Autor:in), Mahmoud Sharabyb (Autor:in), 2020, Genetics Basis of Gonadotropin-Releasing Hormone GnRH, München, GRIN Verlag, https://www.grin.com/document/512579
-
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen. -
Laden Sie Ihre eigenen Arbeiten hoch! Geld verdienen und iPhone X gewinnen.