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Transcriptome analysis of highly purified mouse spermatogenic cell populations: gene expression signatures switch from meiotic-to postmeiotic-related processes at pachytene stage.

da Cruz I, Rodríguez-Casuriaga R, Santiñaque FF, Farías J, Curti G, Capoano CA, Folle GA, Benavente R, Sotelo-Silveira JR, Geisinger A - BMC Genomics (2016)

Bottom Line: Interestingly, we found that a considerable number of genes involved in early as well as late meiotic processes are already on at early meiotic prophase, with a high proportion of them being expressed only for the short time lapse of lepto-zygotene stages.Moreover, we found that a good proportion of the differential gene expression in spermiogenesis corresponds to up-regulation of genes whose expression starts earlier, at pachytene stage; this includes transition protein-and protamine-coding genes, which have long been claimed to switch on during spermiogenesis.In addition, our results afford new insights concerning X chromosome meiotic inactivation and reactivation.

View Article: PubMed Central - PubMed

Affiliation: Department of Genomics, Instituto de Investigaciones Biológicas Clemente Estable (IIBCE), Av. Italia 3318, 11,600, Montevideo, Uruguay.

ABSTRACT

Background: Spermatogenesis is a complex differentiation process that involves the successive and simultaneous execution of three different gene expression programs: mitotic proliferation of spermatogonia, meiosis, and spermiogenesis. Testicular cell heterogeneity has hindered its molecular analyses. Moreover, the characterization of short, poorly represented cell stages such as initial meiotic prophase ones (leptotene and zygotene) has remained elusive, despite their crucial importance for understanding the fundamentals of meiosis.

Results: We have developed a flow cytometry-based approach for obtaining highly pure stage-specific spermatogenic cell populations, including early meiotic prophase. Here we combined this methodology with next generation sequencing, which enabled the analysis of meiotic and postmeiotic gene expression signatures in mouse with unprecedented reliability. Interestingly, we found that a considerable number of genes involved in early as well as late meiotic processes are already on at early meiotic prophase, with a high proportion of them being expressed only for the short time lapse of lepto-zygotene stages. Besides, we observed a massive change in gene expression patterns during medium meiotic prophase (pachytene) when mostly genes related to spermiogenesis and sperm function are already turned on. This indicates that the transcriptional switch from meiosis to post-meiosis takes place very early, during meiotic prophase, thus disclosing a higher incidence of post-transcriptional regulation in spermatogenesis than previously reported. Moreover, we found that a good proportion of the differential gene expression in spermiogenesis corresponds to up-regulation of genes whose expression starts earlier, at pachytene stage; this includes transition protein-and protamine-coding genes, which have long been claimed to switch on during spermiogenesis. In addition, our results afford new insights concerning X chromosome meiotic inactivation and reactivation.

Conclusions: This work provides for the first time an overview of the time course for the massive onset and turning off of the meiotic and spermiogenic genetic programs. Importantly, our data represent a highly reliable information set about gene expression in pure testicular cell populations including early meiotic prophase, for further data mining towards the elucidation of the molecular bases of male reproduction in mammals.

No MeSH data available.


Related in: MedlinePlus

Representation of DEG between pairwise sample comparisons of the four populations in chronological order. The following comparisons were performed: 2C/LZ; LZ/PS; PS/RS (/FC/ ≥ 2; Kal’s test p ≤ 0.01). a. Heat map of expression levels and hierarchical clustering for the global gene expression in the four samples. All genes detected as differential in at least one sample were included. Z-score values are coded on the green-to-red scale (high expression: red; low expression: green). b. Venn diagram of up-regulated and down-regulated genes. Separate and overlapping expression between samples is shown. c. Temporal expression profiles of DEG, ordered based on the p-value significance of the number of assigned vs expected genes. Only the 10 most significant profiles are shown. The p-value (bottom of each panel) and number of genes (below) for each profile are shown
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Fig3: Representation of DEG between pairwise sample comparisons of the four populations in chronological order. The following comparisons were performed: 2C/LZ; LZ/PS; PS/RS (/FC/ ≥ 2; Kal’s test p ≤ 0.01). a. Heat map of expression levels and hierarchical clustering for the global gene expression in the four samples. All genes detected as differential in at least one sample were included. Z-score values are coded on the green-to-red scale (high expression: red; low expression: green). b. Venn diagram of up-regulated and down-regulated genes. Separate and overlapping expression between samples is shown. c. Temporal expression profiles of DEG, ordered based on the p-value significance of the number of assigned vs expected genes. Only the 10 most significant profiles are shown. The p-value (bottom of each panel) and number of genes (below) for each profile are shown

Mentions: Hierarchical clustering of differentially expressed genes (DEG) showed relatively similar patterns between the 2C and LZ populations on one side, and between the PS and RS populations on the other. Moreover, the global pattern of turned on/turned off genes appeared practically reversed between the LZ and PS cell populations (Fig. 3a). Accordingly, the highest number of DEG was found when comparing LZ and PS cell populations, both for up-regulated and for down-regulated genes (Table 1 and Fig. 3b). In addition, as illustrated in Fig. 3c, temporal expression profile analysis of the DEG showed that the highest number of genes under a single profile (n = 668) pertains to genes that are up-regulated between LZ and PS (profile 1).Fig. 3


Transcriptome analysis of highly purified mouse spermatogenic cell populations: gene expression signatures switch from meiotic-to postmeiotic-related processes at pachytene stage.

da Cruz I, Rodríguez-Casuriaga R, Santiñaque FF, Farías J, Curti G, Capoano CA, Folle GA, Benavente R, Sotelo-Silveira JR, Geisinger A - BMC Genomics (2016)

Representation of DEG between pairwise sample comparisons of the four populations in chronological order. The following comparisons were performed: 2C/LZ; LZ/PS; PS/RS (/FC/ ≥ 2; Kal’s test p ≤ 0.01). a. Heat map of expression levels and hierarchical clustering for the global gene expression in the four samples. All genes detected as differential in at least one sample were included. Z-score values are coded on the green-to-red scale (high expression: red; low expression: green). b. Venn diagram of up-regulated and down-regulated genes. Separate and overlapping expression between samples is shown. c. Temporal expression profiles of DEG, ordered based on the p-value significance of the number of assigned vs expected genes. Only the 10 most significant profiles are shown. The p-value (bottom of each panel) and number of genes (below) for each profile are shown
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4837615&req=5

Fig3: Representation of DEG between pairwise sample comparisons of the four populations in chronological order. The following comparisons were performed: 2C/LZ; LZ/PS; PS/RS (/FC/ ≥ 2; Kal’s test p ≤ 0.01). a. Heat map of expression levels and hierarchical clustering for the global gene expression in the four samples. All genes detected as differential in at least one sample were included. Z-score values are coded on the green-to-red scale (high expression: red; low expression: green). b. Venn diagram of up-regulated and down-regulated genes. Separate and overlapping expression between samples is shown. c. Temporal expression profiles of DEG, ordered based on the p-value significance of the number of assigned vs expected genes. Only the 10 most significant profiles are shown. The p-value (bottom of each panel) and number of genes (below) for each profile are shown
Mentions: Hierarchical clustering of differentially expressed genes (DEG) showed relatively similar patterns between the 2C and LZ populations on one side, and between the PS and RS populations on the other. Moreover, the global pattern of turned on/turned off genes appeared practically reversed between the LZ and PS cell populations (Fig. 3a). Accordingly, the highest number of DEG was found when comparing LZ and PS cell populations, both for up-regulated and for down-regulated genes (Table 1 and Fig. 3b). In addition, as illustrated in Fig. 3c, temporal expression profile analysis of the DEG showed that the highest number of genes under a single profile (n = 668) pertains to genes that are up-regulated between LZ and PS (profile 1).Fig. 3

Bottom Line: Interestingly, we found that a considerable number of genes involved in early as well as late meiotic processes are already on at early meiotic prophase, with a high proportion of them being expressed only for the short time lapse of lepto-zygotene stages.Moreover, we found that a good proportion of the differential gene expression in spermiogenesis corresponds to up-regulation of genes whose expression starts earlier, at pachytene stage; this includes transition protein-and protamine-coding genes, which have long been claimed to switch on during spermiogenesis.In addition, our results afford new insights concerning X chromosome meiotic inactivation and reactivation.

View Article: PubMed Central - PubMed

Affiliation: Department of Genomics, Instituto de Investigaciones Biológicas Clemente Estable (IIBCE), Av. Italia 3318, 11,600, Montevideo, Uruguay.

ABSTRACT

Background: Spermatogenesis is a complex differentiation process that involves the successive and simultaneous execution of three different gene expression programs: mitotic proliferation of spermatogonia, meiosis, and spermiogenesis. Testicular cell heterogeneity has hindered its molecular analyses. Moreover, the characterization of short, poorly represented cell stages such as initial meiotic prophase ones (leptotene and zygotene) has remained elusive, despite their crucial importance for understanding the fundamentals of meiosis.

Results: We have developed a flow cytometry-based approach for obtaining highly pure stage-specific spermatogenic cell populations, including early meiotic prophase. Here we combined this methodology with next generation sequencing, which enabled the analysis of meiotic and postmeiotic gene expression signatures in mouse with unprecedented reliability. Interestingly, we found that a considerable number of genes involved in early as well as late meiotic processes are already on at early meiotic prophase, with a high proportion of them being expressed only for the short time lapse of lepto-zygotene stages. Besides, we observed a massive change in gene expression patterns during medium meiotic prophase (pachytene) when mostly genes related to spermiogenesis and sperm function are already turned on. This indicates that the transcriptional switch from meiosis to post-meiosis takes place very early, during meiotic prophase, thus disclosing a higher incidence of post-transcriptional regulation in spermatogenesis than previously reported. Moreover, we found that a good proportion of the differential gene expression in spermiogenesis corresponds to up-regulation of genes whose expression starts earlier, at pachytene stage; this includes transition protein-and protamine-coding genes, which have long been claimed to switch on during spermiogenesis. In addition, our results afford new insights concerning X chromosome meiotic inactivation and reactivation.

Conclusions: This work provides for the first time an overview of the time course for the massive onset and turning off of the meiotic and spermiogenic genetic programs. Importantly, our data represent a highly reliable information set about gene expression in pure testicular cell populations including early meiotic prophase, for further data mining towards the elucidation of the molecular bases of male reproduction in mammals.

No MeSH data available.


Related in: MedlinePlus