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Developmental regulation of Rtdpoz-T1 and -T2. (A) Exons constituting the testicular T1 and T2 transcripts. In both T1 and T2, the common exon 1a is used. The noncoding sequences are shown as unfilled boxes. The uninterrupted T1- and T2-coding sequences that reside in exons 4 (T1–4) and 3 (T2–3) of the respective genes are shown as cross-hatched or slanting-hatched boxes, respectively. In T1, the L1 sequence (see text) in the 3'-UTR is shown in grey. The relative positions of the primers used in the RT-PCR expression profiling and the RACE analysis are shown (see Table 2 for primer sequences). An, polyA tract. (B) Developmental expression profiles of the T1 and T2 genes. The developmental stages analysed were from day 12 (E12) through to day 20 (E20). β-actin was included as a control. In the experiments, the Ex1a-B + T1SP-R1 or Ex1a-B + T2SP-R1 primer pairs (see above) were used in the first-round PCR for T1 and T2, respectively, followed by the use of the Ex1a-A + T1SP-R3 (for T1) or Ex1a-A + T2SP-R3 (for T2) primer pairs in the second-round PCR as detailed in the Methods section. On the left and right of the photo panels are schematic representations of the PCR bands with band designations as explained in the text. (C) 3'-Extended RT-PCR analysis of the T1 transcripts in the developmental stages that expressed the gene. In the first-round PCR, the Ex1a-B and 3096R primers (see (A) above) were used followed by a second-round PCR using the Ex1a-A and 2965R primers as described in Methods. Band designations, prefixed by "XT", are depicted in the schematic drawing alongside the gel display.
Source publication
Retrotransposition is an important evolutionary force for the creation of new and potentially functional intronless genes which are collectively called retrogenes. Many retrogenes are expressed in the testis and the gene products have been shown to actively participate in spermatogenesis and other unique functions of the male germline. We have prev...
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Citations
... The rtdpoz-T1 and rtdpoz-T2 retrogenes, specifically expressed in testis and in the developing embryo in rat, and supposed to encode nuclear scaffold proteins functioning as transcription regulators, have multiple exons deriving from TE sequences [58,59]. For example, rtdpoz-T1 has 5 out of 8 exons and an alternative polyadenylation signal that are derived from various TEs, mainly L1 and ERVs. ...
... For example, rtdpoz-T1 has 5 out of 8 exons and an alternative polyadenylation signal that are derived from various TEs, mainly L1 and ERVs. These TE-derived exons may be implicated in the translational regulation of these transcripts, notably through the formation of upstream ORFs [59]. ...
Transposable elements (TEs) are major components of all vertebrate genomes that can cause deleterious insertions and genomic instability. However, depending on the specific genomic context of their insertion site, TE sequences can sometimes get positively selected, leading to what are called “exaptation” events. TE sequence exaptation constitutes an important source of novelties for gene, genome and organism evolution, giving rise to new regulatory sequences, protein-coding exons/genes and non-coding RNAs, which can play various roles beneficial to the host. In this review, we focus on the development of vertebrates, which present many derived traits such as bones, adaptive immunity and a complex brain. We illustrate how TE-derived sequences have given rise to developmental innovations in vertebrates and how they thereby contributed to the evolutionary success of this lineage.
... TEs have often been considered as parasites, or background genomic noise, but they are increasingly appreciated for their expanding number of roles in genome evolution, and gene regulation rewiring [139][140][141]. Importantly, TEs are a major contributor to the sequences of noncoding RNAs [119] and several studies have observed the splicing of distal TE exons into coding sequences that did not contain any TEs [142][143][144]. For example, MERVL expression marks a subpopulation of totipotent cells in cultures of embryonic stem cells [145,146], and a distinctive feature of these cells is the splicing of MERVLs into coding genes, such as Zfp352 and Apol7b [147]. ...
The measurement of gene expression has long provided significant insight into biological functions. The development of high-throughput short-read sequencing technology has revealed transcriptional complexity at an unprecedented scale, and informed almost all areas of biology. However, as researchers have sought to gather more insights from the data, these new technologies have also increased the computational analysis burden. In this review, we describe typical computational pipelines for RNA-Seq analysis and discuss their strengths and weaknesses for the assembly, quantification and analysis of coding and non-coding RNAs. We also discuss the assembly of transposable elements into transcripts, and the difficulty these repetitive elements pose. In summary, RNA-Seq is a powerful technology that is likely to remain a key asset in the biologist's toolkit.
... For comparison, the minimal prokaryotic genome size in the Micoplasma genitalium is less than the maximal in the Myxococcus xanthus by just 16 times (Patrushev and Minkevich, 2007). Despite the fact that the portion of NSs encoding the proteins in eukaryotic genomes is from a part to several percent of all DNA, TEs were the most important sources of their emergence and evolution by duplicating the existing genes (Cerbin and Jiang, 2018;Grandi et al., 2015;Huang et al., 2009;Kubiak and Makalowska, 2017;Sakai et al., 2011;Tan et al., 2016;Zhu et al., 2016), exonization (Elkon et al., 2013;Tajnik et al., 2015;Wang et al., 2012), and domestication of TEs themselves (Alzohairy et al., 2013;Duan et al., 2017;Feschotte, 2008;Garavis et al., 2013;Schrader and Schmitz, 2018;Sinzelle et al., 2009;Volff et al., 2006;Zdobnov et al., 2005). ...
... It is assumed that the number of retrogens is much greater than that determined by standard identification methods due to 5'-truncation, although approximately 10% of all PCGs contain one and more retrocopies even taking this into account (Grandi et al., 2015). For example, more than 1000 retrogens are contained in the human genome that are transcribed and show biological activity (Huang et al., 2009). In rice, the expression of at least 66% of detected retrocopies was determined; however, they were with lower values than their initial genes but with tissue-specific correlation of their levels (Sakai et al., 2011). ...
The ability of transposons to unite genes separated by their insertions encoding common biological processes into regulatory networks contributed, simultaneously with the complication of eukaryotes, to their evolutionary success by forming new universal systems. By means of these systems, including DNA methylation, histone modifications, the relationship of telomeres with transposons, splicing regulation, and RNA interference, global distribution of transposons in the genomes was accompanied by the emergence of their structural innovations, dynamic regulatory sequences, and protein-coding genes. The mobile elements contributed to the evolution of protein-coding genes by their duplication, as well as exonization, and domestication of the transposons themselves. The resulting new genes contain transposon sequences involved in their management by means of regulatory networks and noncoding RNAs also originating from the mobile elements. A strategy wherein the translation of noncoding RNA genes contributed to the selection of the obtained polypeptides as functional cellular proteins was developed during evolution. At the same time, noncoding RNAs are also processed into molecules involved in the regulatory processes independently or as a part of the protein complexes. The duality of functions was inherent to all noncoding RNAs whose nonrandom decay/processing leads to the formation of molecules that have a regulatory effect on the transposons and protein-coding genes. A strategy wherein primary transposon transcripts interact with different systems of their processing (arisen to protect the hosts from transposons), forming functional RNA molecules translated into the peptides, was developed in the evolution of eukaryotes. The transposons are universal sources for these strategies; this explains their global distribution in eukaryotic genomes and domestication in the system of “double search” for targets for functional interaction of noncoding RNAs and processed products of their translation. In addition to splicing, primary transcripts of some protein-coding genes can also be processed in functional noncoding RNAs involved in common biological reactions with the gene protein product. This substantiates the associations of multifactorial diseases with the gene SNP since they can cause inactivation of RNA domains. It was suggested that functional dualism of the transposon transcripts could be an important condition of the emergence of life, while the mobile elements are one of fundamental properties of living.
... These interactions are fundamental for programmed cell differentiation [29]. Presumably, the same principle grounds the evolutionary genesis of regulatory pathways of the expression of protein-encoding genes, in the origination of which transposons had a great impact in evolution via ME exonization [11,46] and the domestication of MEs [11,13,20,21,35,53] or their parts during gene duplication [26,29]. ...
... Apart from introns, intergenic MEs are also used in exonization, along with the creation of alternative 3'UTR [46]. Retroelements promote gene duplication via the formation of retrocopies, with many of them becoming functional [26]. Over 10000 pseudogenes originating from the duplication of protein-encoding genes have accumulated in the human genome [29], and at least 1000 of them transcribe [26]. ...
... Retroelements promote gene duplication via the formation of retrocopies, with many of them becoming functional [26]. Over 10000 pseudogenes originating from the duplication of protein-encoding genes have accumulated in the human genome [29], and at least 1000 of them transcribe [26]. ...
In regulation of gene expression in the ontogenesis of multicellular eukaryotes, in addition to transcription factors, an important role is played by epigenetic factors that control the release of genetic information in each cell division. Many binding sites for the transcription factors were derived from transposons sequences. Mobile elements are also important sources of non-coding RNA. Due to this, transposons have an indirect effect on gene expression and genome methylation. In evolution, transposons serve as important sources for the origin of new protein and proteins domains. A number of studies have identified that long non-coding RNAs and microRNAs can be translated into functional peptides. At the same time, transposons remain active in the hypothalamus of adult humans, which is consistent with the transcription of non-coding RNAs in these structures, which may be key in aging.
... Retrotransposed copies lack many of their parental gene genetic features, such as introns and regulatory elements (Pan and Zhang 2009). Retrotransposition is an important evolutionary force for the origin of new and potentially functional retrogenes, which are intron-depleted (Huang et al. 2009). Retrocopies of hnRNP Q-like were not detected in the cichlid P. nyererei, which also harbors a B chromosome (Valente et al. 2014), or in any other cichlid genome investigated here. ...
B chromosomes are dispensable elements observed in many eukaryotic species, including the African cichlid Astatotilapia latifasciata, which might have one or two B chromosomes. Although there have been many studies focused on the biology of these chromosomes, questions about the evolution, maintenance, and potential effects of these chromosomes remain. Here, we identified a variant form of the hnRNP Q-like gene inserted into the B chromosome of A. latifasciata that is characterized by a high copy number and intron-less structure. The absence of introns and presence of transposable elements with a reverse transcriptase domain flanking hnRNP Q-like sequences suggest that this gene was retroinserted into the B chromosome. RNA-Seq analysis did not show that the B variant retroinserted copies are transcriptionally active. However, RT-qPCR results showed variations in the canonical hnRNP Q-like copy expression levels among exons, tissues, sex, and B presence/absence. Although the patterns of transcription are not well understood, the exons of the B retrocopies were overexpressed, and a bias for female B+ expression was also observed. These results suggest that retroinsertion is an additional and important mechanism contributing to B chromosome formation. Furthermore, these findings indicate a bias towards female differential expression of B chromosome sequences, suggesting that B chromosomes and sex determination are somehow associated in cichlids.
... It has been demonstrated that L1 can affect transcription in several distinct ways. They can generate alternative splice sites resulting in the exonization of L1 sequences, at least in rodents (Zemojtel et al. 2007;Huang et al. 2009). Also intronic L1s may sometimes interfere with transcriptional elongation and so produce different lengths of mRNA from a gene (Han et al. 2004). ...
Retrotransposons comprise over 40 % of the human genome and are a major contributor to genome diversity and evolution. They contribute to human genome variation through both germline and somatic retrotransposition. Over recent years, studies on the biology of cancer have revealed that somatic retrotransposition is a feature of many cancer genomes. The most recent comparison between 200 pairs of tumours and normal tissue, across 11 tumour types, has revealed frequent somatic retrotransposition in particular cancers; lung squamous cell, head and neck squamous cell, colorectal and endometrial carcinomas. Importantly some of these insertions occur in cancer-related genes underlining retrotransposition’s role as a mutagen. It is now clear that retrotransposons contribute to genome instability during cancer progression. However, the exact role of retrotransposons in tumuorigenesis, tumour progression and prognosis still remains a subject of an active discussion in the field of cancer biology.
In this chapter, we have attempted to explain the biology of retrotransposons in the human genome, with the main focus on LINE-1 elements. We then have discussed how LINE-1 causes genome instability in the genome and the host defence mechanisms deployed to supress their retrotransposition. Next, we discuss the role of LINE-1 activity during tumourigenesis and consider the recent findings concerning their activity in different types of cancers. Finally, we explore how retrotransposons can be used as diagnostic tools in cancer.
... reading frames (uORFs), for translational regulation [21][22][23]. We also found that testes-enriched novel 5′-UTRs have abundant uORFs (n = 56, 50%) with some of 197 novel exons in the testes, suggesting a testes-specific regulatory role in translation. ...
Background
RNA sequencing (RNA-seq) has revolutionized the detection of transcriptomic signatures due to its high-throughput sequencing ability. Therefore, genomic annotations on different animal species have been rapidly updated using information from tissue-enriched novel transcripts and novel exons.
Results
34 putative novel transcripts and 236 putative tissue-enriched exons were identified using RNA-Seq datasets representing six tissues available in mouse databases. RT-PCR results indicated that expression of 21 and 2 novel transcripts were enriched in testes and liver, respectively, while 31 of the 39 selected novel exons were detected in the testes or heart. The novel isoforms containing the identified novel exons exhibited more dominant expression than the known isoforms in heart and testes. We also identified an example of pathology-associated exclusion of heart-enriched novel exons such as Sorbs1 and Cluh during pressure-overload cardiac hypertrophy.
Conclusion
The present study depicted tissue-enriched novel transcripts, a tissue-specific isoform switch, and pathology-associated alternative splicing in a mouse model, suggesting tissue-specific genomic diversity and plasticity.
Electronic supplementary material
The online version of this article (doi:10.1186/1471-2164-15-592) contains supplementary material, which is available to authorized users.
... Preparation of total RNA and reverse transcription (RT) were performed as previously described (Huang et al. 2009). The relative SPOP mRNA levels were quantified by real-time RT-PCR using the DyNAmo TM Flash SYBR® Green qPCR kit (Finnzymes, Espoo, Finland) as described (Choo et al. 2011). ...
The speckle POZ protein, SPOP, is an adaptor of the Cul3-based ubiquitination process, and has been implicated in the carcinogenesis process. Despite recent elucidation of biological functions, regulation of SPOP gene expression has not been reported. In this study, the mRNA levels of the mouse SPOP (mSPOP) gene were first shown to vary noticeably in different tissues. However, the SPOP protein was detected in high abundance only in Purkinje cells of the cerebellum and seminiferous tubule of the testis, echoing previous reports of involvement of ubiquitination in neuron cells and in spermatogenesis. In other mouse tissues and human cancer cell lines analysed, only low SPOP protein levels were detected. The 3'-untranslated regions of both the mSPOP and human SPOP transcripts harbor a conserved putative miR-145 binding site (BS). In some tissues and cell lines, miR-145 and SPOP protein levels were in an inverse relationship suggesting miR-145 regulation. Luciferase assays of deletion and point mutation constructs of the miR-145 BS, and miR-145 induction by serum starvation that resulted in reduced endogenous SPOP levels provided further evidence that miR-145 is likely involved in post-transcriptional regulation of SPOP expression in selected tissues, and possibly with the participation of other miRNA species.
... PPP1R2 forms a stable and high affinity complex with PPP1C by blocking the active site. The reactivation of the complex is triggered by phosphorylation at Thr72 of PPP1R2 through several kinases, including glycogen synthase 3 (GSK3) [52][53][54]. PPP1R2 is also phosphorylated at the residue Ser86 by casein kinase 2 (CK2) that accelerates the subsequent phosphorylation at Thr72 by GSK3 [16]. The comparison of human PPP1R2P1, PPP1R2P3 and PPP1R2P9 with PPP1R2 amino acid sequences (Figure 7) shows that PPP1R2P9 is the most divergent (41%) and PPP1R2P3 the most similar (95%). ...
... Recently, a report identified the PPP1R2 protein in heat-stable extracts of bull testis and mouse testis and sperm where it may account for this PPP1R2-like activity [58]. It is well known that testis is one of the organs where most pseudogenes are expressed and their gene products were shown to have important roles in spermatogenesis and other germ cell related functions [52][53][54]. This might be due, in part, to the hypertranscription state of the autosomal chromosomes in the meiotic and post-meiotic germ cells due to chromatin modifications [13,54,59]. ...
... It is well known that testis is one of the organs where most pseudogenes are expressed and their gene products were shown to have important roles in spermatogenesis and other germ cell related functions [52][53][54]. This might be due, in part, to the hypertranscription state of the autosomal chromosomes in the meiotic and post-meiotic germ cells due to chromatin modifications [13,54,59]. A recent study done by GENCODE has revealed that 64% of all validated expressed pseudogenes are expressed in testis [3]. ...
Pseudogenes are traditionally considered "dead" genes, therefore lacking biological functions. This view has however been challenged during the last decade. This is the case of the Protein phosphatase 1 regulatory subunit 2 (PPP1R2) or inhibitor-2 gene family, for which several incomplete copies exist scattered throughout the genome.
In this study, the pseudogenization process of PPP1R2 was analyzed. Ten PPP1R2-related pseudogenes (PPP1R2P1-P10), highly similar to PPP1R2, were retrieved from the human genome assembly present in the databases. The phylogenetic analysis of mammalian PPP1R2 and related pseudogenes suggested that PPP1R2P7 and PPP1R2P9 retroposons appeared before the great mammalian radiation, while the remaining pseudogenes are primate-specific and retroposed at different times during Primate evolution. Although considered inactive, four of these pseudogenes seem to be transcribed and possibly possess biological functions. Given the role of PPP1R2 in sperm motility, the presence of these proteins was assessed in human sperm, and two PPP1R2-related proteins were detected, PPP1R2P3 and PPP1R2P9. Signatures of negative and positive selection were also detected in PPP1R2P9, further suggesting a role as a functional protein.
The results show that contrary to initial observations PPP1R2-related pseudogenes are not simple bystanders of the evolutionary process but may rather be at the origin of genes with novel functions.
... Pseudogenes originate from retrotransposition activity, so they have truncated 5'UTR due to the low processivity of the reverse transcriptase, and direct repeats at both ends [38]. Testis is an organ where many pseudogenes are expressed as mRNA and proteins that have been shown to actively participate in spermatogenesis or other germ cell functions [39,40]. Transcription in testis, when compared to other somatic tissues, tends to activate alternative promoters, which are otherwise imperfect or weak [40,41]. ...
... Testis is an organ where many pseudogenes are expressed as mRNA and proteins that have been shown to actively participate in spermatogenesis or other germ cell functions [39,40]. Transcription in testis, when compared to other somatic tissues, tends to activate alternative promoters, which are otherwise imperfect or weak [40,41]. The apparent function of pseudogenes in testis germ cells could be a way to facilitate the appearance of new genes from the parental ones [39]. ...
Background
Protein Ser/Thr Phosphatase PPP1CC2 is an alternatively spliced isoform of PPP1C that is highly enriched in testis and selectively expressed in sperm. Addition of the phosphatase inhibitor toxins okadaic acid or calyculin A to caput and caudal sperm triggers and stimulates motility, respectively. Thus, the endogenous mechanisms of phosphatase inhibition are fundamental for controlling sperm function and should be characterized. Preliminary results have shown a protein phosphatase inhibitor activity resembling PPP1R2 in bovine and primate spermatozoa.
Results
Here we show conclusively, for the first time, that PPP1R2 is present in sperm. In addition, we have also identified a novel protein, PPP1R2P3. The latter was previously thought to be an intron-less pseudogene. We show that the protein corresponding to the pseudogene is expressed. It has PPP1 inhibitory potency similar to PPP1R2. The potential phosphosites in PPP1R2 are substituted by non-phosphorylable residues, T73P and S87R, in PPP1R2P3. We also confirm that PPP1R2/PPP1R2P3 are phosphorylated at Ser121 and Ser122, and report a novel phosphorylation site, Ser127. Subfractionation of sperm structures show that PPP1CC2, PPP1R2/PPP1R2P3 are located in the head and tail structures.
Conclusions
The conclusive identification and localization of sperm PPP1R2 and PPP1R2P3 lays the basis for future studies on their roles in acrosome reaction, sperm motility and hyperactivation. An intriguing possibility is that a switch in PPP1CC2 inhibitory subunits could be the trigger for sperm motility in the epididymis and/or sperm hyperactivation in the female reproductive tract.
