The ovary-specific proteome
The main function of the ovary, a paired organ forming a major component of the female genital system, is to produce and deliver mature oocytes and produce hormones needed for female reproduction. There are approximately 40000 primary oocytes (germ cells), arrested in prophase, at female puberty and from these only some 400 undergo complete meiosis and develop into secondary oocytes. The main constituents of adult ovaries are hormone producing ovarian stromal cells. The transcriptome analysis shows that 64% of all human proteins (n=20344) are expressed in the ovary and 138 of these genes show an elevated expression in ovary compared to other tissue types.
An analysis of the genes with elevated expression in the ovary with regard to tissue distribution shows that these proteins are mainly expressed in ovarian stroma cells and as germ cells are exceedingly rare, follicle or oocyte specific proteins are not detected.
- 6 ovary enriched genes
- 138 genes defined as elevated in the ovary
- A majority of the elevated genes encode proteins expressed in stroma cells
- Most group enriched genes share expression with cerebral cortex and testis
Figure 1. The distribution of all genes across the five categories based on transcript abundance in ovary as well as in all other tissues.
138 genes show some level of elevated expression in the ovary compared to other tissues. The four categories of genes with elevated expression in ovary compared to other organs are shown in Table 1.
Table 1. The genes with elevated expression in ovary
Number of genes
||At least five-fold higher mRNA levels in a particular tissue as compared to all other tissues
||At least five-fold higher mRNA levels in a group of 2-7 tissues
||At least five-fold higher mRNA levels in a particular tissue as compared to average levels in all tissues
||Total number of elevated genes in ovary
Table 2. The 6 genes with the highest level of enriched expression in ovary. "Predicted localization" shows the classification of each gene into three main classes: Secreted, Membrane, and Intracellular, where the latter consists of genes without any predicted membrane and secreted features. "mRNA (tissue)" shows the transcript level as FPKM values, TS-score (Tissue Specificity score) corresponds to the score calculated as the fold change to the second highest tissue.
||WAP, follistatin/kazal, immunoglobulin, kunitz and netrin domain containing 2
||endothelin converting enzyme-like 1
||kelch domain containing 8A
||crystallin, gamma D
||melanoma associated antigen (mutated) 1-like 1
Some of the proteins predicted to be membrane-spanning are intracellular, e.g., in the Golgi or mitochondrial membranes, and some of the proteins predicted to be secreted can potentially be retained in a compartment belonging to the secretory pathway, such as the ER, or remain attached to the outer face of the cell membrane by a GPI anchor.
The ovary transcriptome
An analysis of the expression levels of each gene makes it possible to calculate the relative mRNA pool for each of the categories. The analysis shows that 90% of the mRNA molecules derived from ovary correspond to housekeeping genes and only 2% of the mRNA pool corresponds to genes categorized as ovary enriched, group enriched, or ovary enhanced. Thus, most of the transcriptional activity in the ovary relates to proteins with presumed housekeeping functions as they are found in all tissues and cells analyzed.
Protein expression of genes elevated in ovary
In-depth analysis of the elevated genes in ovary using antibody-based protein profiling allowed us to visualize where these proteins are expressed within the ovary including follicle cells and stroma cells.
Examples of proteins with elevated expression in the ovary include MUM1L1, a putative protein containing a mutated melanoma-associated antigen 1 domain, expressed both in ovarian stroma cells and follicle cells, RBP1, a carrier protein involved in the transport of retinol, COL16A1, the alpha chain of type XVI collagen, and CDH11, a type II classical cadherin that mediates calcium-dependent cell-cell adhesion all expressed in stroma cells.
Genes shared between ovary and other tissues
There are 26 group-enriched genes expressed in the ovary. Group enriched genes are defined as genes showing a 5-fold higher average level of mRNA expression in a group of 2-7 tissues, including ovary, compared to all other tissues.
In order to illustrate the relation of ovarian tissue to other tissue types, a network plot was generated, displaying the number of commonly expressed genes between different tissue types.
Figure 2. An interactive network plot of the ovary enriched and group enriched genes connected to their respective enriched tissues (grey circles). Red nodes represent the number of ovary enriched genes and orange nodes represent the number of genes that are group enriched. The sizes of the red and orange nodes are related to the number of genes displayed within the node. Each node is clickable and results in a list of all enriched genes connected to the highlighted edges. The network is limited to group enriched genes in combinations of up to 3 tissues, but the resulting lists show the complete set of group enriched genes in the particular tissue.
The ovary shares of a few genes with several other tissue types but has most shared group enriched genes with cerebral cortex (10 genes) and testis (8 genes). MAG, a myelin-associated glycoprotein, is one example of a group enriched gene shared between ovary and cerebral cortex.
RGS22, a putative regulator of G-protein signaling with evidence only at transcript level, is one example of proteins expressed in testis, fallopian tube and ovary and PEG3, a paternally expressed zinc finger protein, shows enhanced expression in ovary and placenta.
The function of the ovary is to develop female germinal cells, oocytes, and to produce hormones estrogen, testosterone and progesterone which are necessary for reproduction. Estrogen is responsible for the appearance of secondary sex characteristics for females at puberty and for the maturation and maintenance of the reproductive organs in their mature functional state. Progesterone prepares the uterus for pregnancy and the mammary glands for lactation. Together with estrogen, progesterone functions by promoting menstrual cycle changes in the endometrium.
The ovarian follicle is an anatomical structure in which the primary oocyte develops. The cells types of the ovarian follicle include the oocyte and the granulosa cells which surround the follicle. The granulosa cells, in turn, are enclosed in a thin layer of extracellular matrix - the follicular basement membrane or basal lamina. Outside the basal lamina, the layers theca interna and theca externa are found.
In the reproductive period the primordial follicles are scattered irregularly in clusters throughout the superficial cortex of the ovary. The primordial follicles eventually develop into primary, secondary and tertiary vesicular follicles. Each month, typically only one developing primary follicle becomes dominant, and achieves complete maturation to release the oocyte. Other developing follicles undergo atresia.
The follicular phase lasts from the beginning of menstruation to the start of ovulation and is the phase of the menstrual cycle during which the ovarian follicles mature.
Follicle-stimulating hormone (FSH), produced by the anterior pituitary gland, begins to rise in the last few days of the previous menstrual cycle and is highest during the first week of the follicular phase. The rise in FSH levels recruits tertiary-stage ovarian follicles for entry into the menstrual cycle.
FSH induces the proliferation of and expression of luteinizing hormone (LH) receptors on the granulosa cells in the developing follicles. Under the influence of FSH the granulosa cells begins to secrete estrogen. The increase of estrogen stimulates production of gonadotropin-releasing hormone (GnRH), by the hypothalamus, which in turn increases production of LH, by the anterior pituitary gland. LH induces androgen synthesis by thecal cells, stimulates proliferation, differentiation, and secretion of follicular thecal cells and increases LH receptor expression on granulosa cells.
Two or three days before LH levels begin to increase normally one of the follicles has emerged as dominant. The increase in estrogen levels initiate the formation of a new layer of endometrium in the uterus, histologically identified as the proliferative endometrium.
Estrogen levels peak towards the end of the follicular phase, which causes a surge in levels of LH and FSH. This lasts from 24 to 36 hours, and results in the rupture of the ovarian follicles, causing the oocyte to be released from the ovary via the fallopian tube. The oocyte, or ovum, will travel down one of the fallopian tubes, pushed along by cilia, beginning its journey toward the uterus.
If no fertilization occurs, the ovum will degenerate between 12 and 24 hours after ovulation and eventually be discharged through menstruation.
Figure 3. The ovaries, fallopian tubes and uterus. The menstrual cycle can be described both by the uterine cycle and the ovarian cycle which consists of the follicular phase, ovulation, and the luteal phase. During the follicular phase follicles in the ovary start to mature and during ovulation the dominant follicle releases an ovum (egg) via the fallopian tube. After ovulation, during the luteal phase, the remaining parts of the follicle transforms into the corpus luteum which produces hormones that support the early pregnancy or if the ovum is not fertilized will atrophy leading to falling levels of hormones and the beginning of the next menstrual cycle. If fertilized the ovum develops into a blastocyst and implantation in the uterus is made possible through changes in both the blastocyst and endometrial wall. Implantation allows for the next step in embryogenesis which includes formation of the placenta.
After ovulation FSH and LH cause the remaining parts of the dominant follicle to transform into the corpus luteum. It continues to grow for some time after ovulation and produces significant amounts of hormones, particularly progesterone and, to a lesser extent, estrogen. Progesterone plays a vital role in making the endometrium receptive to implantation of the blastocyst and supportive of the early pregnancy.
The hormones produced by the corpus luteum suppress the production of FSH and LH that the corpus luteum needs to maintain itself. With continued low levels of FSH and LH, the corpus luteum will atrophy. The death of the corpus luteum results in falling levels of progesterone and estrogen. These falling levels of ovarian hormones cause increased levels of FSH, which begins recruiting follicles for the next cycle. Continued drops in levels of estrogen and progesterone trigger the end of the luteal phase: menstruation and the beginning of the next cycle.
The loss of the corpus luteum can be prevented by implantation of an embryo: after implantation, human embryos produce human chorionic gonadotropin (hCG), which is structurally similar to LH and can preserve the corpus luteum. If implantation occurs, the corpus luteum will continue to produce progesterone for eight to twelve weeks, after which the placenta takes over this function.
The ovaries are paired organs that lie on either side of the uterus close to the lateral pelvic wall, behind the broad ligament and anterior to the rectum. Each ovary is attached to a broad ligament along by a double fold of peritoneum, the mesovarium.
Ovaries of adult women in the reproductive age-group are approximately 3 to 5 x 1.5 to 3 x 0.6-1.5 cm but their size varies considerably due to follicular derivatives. The external surface is usually convoluted. Three ill-defined zones may be discerned on the cut surface: an outer cortex, a central medulla and the hilus. Follicular structures (cystic follicles, corpora lutea, white corpora albicantia) are usually visible in the cortex.
The ovarian cortex consists of ovarian follicles with stroma in between them. Included in the follicles are the cumulus oophorus, membrana granulosa (and the granulosa cells inside it), corona radiata, zona pellucida, and primary oocyte. The zona pellucida, theca of follicle, antrum and liquor folliculi are also contained in the follicle. Also in the cortex is the corpus luteum derived from the follicles.
The innermost layer is the ovarian medulla. It can be hard to distinguish between the cortex and medulla, but follicles are usually not found in the medulla.
The ovary also contains blood vessels and lymphatics.
The surface epithelium, or germinal epithelium, of the ovary forms a simple, focally pseudostratified layer. The cells vary from flat to cuboidal to columnar, and several types may be seen in different areas of the same ovary. The cells are separated from the underlying stroma by a basement membrane.
The primordial follicles are comprised of a primary oocyte, 40 to 70 µm in diameter, surrounded by a single layer of mitotically inactive squamous follicular cells, resting on thin basal lamina.
The histology of human ovary including detailed images and information about the different cell types can be viewed in the Protein Atlas Histology Dictionary.
Here, the protein-coding genes expressed in the ovary are described and characterized, together with examples of immunohistochemically stained tissue sections that visualize protein expression patterns of proteins that correspond to genes with elevated expression in the ovary.
Transcript profiling and RNA-data analyses based on normal human tissues have been described previously (Fagerberg et al., 2013). Analyses of mRNA expression including over 99% of all human protein-coding genes was performed using deep RNA sequencing of 122 individual samples corresponding to 32 different human normal tissue types. RNA sequencing results of 3 fresh frozen tissues representing normal ovary was compared to 119 other tissue samples corresponding to 31 tissue types, in order to determine genes with elevated expression in ovary. A tissue-specific score, defined as the ratio between mRNA levels in ovary compared to the mRNA levels in all other tissues, was used to divide the genes into different categories of expression.
These categories include: genes with elevated expression in ovary, genes expressed in all tissues, genes with a mixed expression pattern, genes not expressed in ovary, and genes not expressed in any tissue. Genes with elevated expression in ovary were further sub-categorized as i) genes with enriched expression in ovary, ii) genes with group enriched expression including ovary and iii) genes with enhanced expression in ovary.
Human tissue samples used for protein and mRNA expression analyses were collected and handled in accordance with Swedish laws and regulation and obtained from the Department of Pathology, Uppsala University Hospital, Uppsala, Sweden as part of the sample collection governed by the Uppsala Biobank. All human tissue samples used in the present study were anonymized in accordance with approval and advisory report from the Uppsala Ethical Review Board.
Relevant links and publications
Uhlen et al (2015). Tissue-based map of the human proteome. Science.
Analysis of the Human Tissue-specific Expression by Genome-wide Integration of Transcriptomics and Antibody-based Proteomics
Histology dictionary - the ovary