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Cold Spring Harb Perspect Biol. Author manuscript; available in PMC Jun 13, 2013.
Published in final edited form as:
PMCID: PMC3428765
NIHMSID: NIHMS461409
Transcriptional Networks in Liver and Intestinal Development
Karyn L. Sheaffer and Klaus H. Kaestnercorresponding author
Department of Genetics, Institute for Diabetes, Obesity, and Metabolism, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, 19104, Phone: 215-898-8759, Fax: 215-573-5892
corresponding authorCorresponding author.
Karyn L. Sheaffer: karyns/at/mail.med.upenn.edu; Klaus H. Kaestner: kaestner/at/mail.med.upenn.edu
Development of the gastrointestinal tract is a complex process that uses unique mechanisms and depends heavily on transcriptional activation. Here we discuss the regionalization of the primitive gut and formation of the intestine and liver. Anterior-posterior positioning in the primitive gut is important for establishing regions that will become functional organs. Coordination of transcription factors and signaling factors between the epithelium and mesenchyme is required for intestinal development and homeostasis. Liver development utilizes a complex transcriptional network that is required for organ domain establishment, cell differentiation, and adult function. Discussion of these transcriptional mechanisms gives us insight on how the primitive gut, composed of simple endodermal cells, develops into multiple diverse cell types that are organized into very complex functional mature organs.
The development of the gastrointestinal tract is a complex and unique process. The gut, unlike other organ systems, is composed of multiple specialized cell types with contributions from three germ layers. The endoderm forms the epithelium of the stomach, intestine, lung, liver, and pancreas. The mesoderm forms both striated (in the esophagus) and smooth muscle that is responsible for peristaltic movements. The neural crest is critical for the enteric nervous system, which controls peristalsis and which is absolutely essential for the proper functioning of the digestive system.
During gastrulation as the endoderm, mesoderm and ectoderm are specified the primitive gut divided into regions with distinct gene expression patterns along the anterior-posterior (AP) axis. These regions are important because each derivative organ is dependent on very diverse developmental programs and to achieve its unique adult function. As cells divide, correct cell positioning within the embryo is required for signaling across germ layers. The interplay between multiple signals and transcription factors is critical in both time and space for correct development. Thus, the primitive gut is divided into foregut, midgut and hindgut. The foregut forms the esophagus, lungs, thyroid, stomach, liver and pancreas. The midgut and hindgut form the small and large intestine (colon), respectively. There are three different mechanisms that are continuously used throughout gut development to maintain regional identity (Figure 1).
Figure 1
Figure 1
Molecular mechanisms guiding gut development
The first is the use of combinations of transcription factors for tight coordination of gene expression in both time and space (Figure 1A). Master transcriptional regulators are required for both the initial specification of the endoderm as well as the appropriate coordination of downstream factors important for different stages of differentiation. Often, it is combinations of several transcription factors, rather than simple factors, that are required to activate the appropriate downstream gene expression programs. This combinational control helps to fine tune activation of particular sets of genes that execute the final function of the derivate organs.
Second, extracellular signaling factors are required at various times throughout intestinal development (Figure 1B). Interestingly, the same signaling factor may cause different and even opposing downstream effects at different times in ontogeny. For instance, a signal factor that is required early for repression of foregut genes must act later for expression to activate the same genes. A good example of this are the fibroblast growth factors (FGF) and bone morphogenetic proteins (BMP), which represses liver-specific genes in the foregut and later are required in the liver primordium for differentiation.
Third, cell positioning and morphogenesis are critical for appropriate signaling between neighboring tissues during development and homeostasis (Figure 1C). Establishment of the subdivisions of the primitive gut requires the activation of specific transcription factors within the endoderm. Activation of many of these factors requires a signaling input from neighboring tissues, especially the mesoderm. As an example, Wnt signaling in the mesoderm is required for anterior-posterior patterning early in development as well as the maintenance of intestinal progenitor proliferation later in the adult.
1. Initial establishment of regional identities
The primitive streak is the first morphological sign of anterior-posterior (AP) positioning in the vertebrate embryo. The process of the establishment of the primitive streak has been reviewed in detail elsewhere (Rivera-Perez 2007; Lee and Anderson 2008). Primitive streak cells form the progenitors for the three germ layers, endoderm, mesoderm, and ectoderm. Early, Mixl1, a member of the Mix/Bix family of paired-like homeodomain proteins, is essential for the establishment of Nodal signaling within the primitive streak (Hart et al. 2002). Subsequently Nodal, a transforming growth factor beta (TGF-β) family member, is required for the activation of multiple transcription factors that function in endoderm specification such as Sox17, FoxA2 and Hhex (Shen 2007; Zorn and Wells 2007).
During gastrulation, movement out of the primitive streak causes early anterior-posterior regionalization of the gut. Using transgenic techniques, endodermal cells have been traced from gastrulation to early organogenesis using fluorescent markers (Tam et al. 2007; Franklin et al. 2008). Cells that leave the primitive streak first are specified as anterior endoderm, while cells migrating later form the posterior endoderm (Lawson et al. 1986; Lawson and Pedersen 1987; Tam and Beddington 1992). These different groups of cells form anterior and posterior pockets of endoderm, also termed the anterior and posterior intestinal portals, respectively. These pockets then elongate towards both ends of the embryo, while the intervening sheet of endoderm closes ventrally to form a connected tube. This process requires Wnt signaling in many organisms, although the situation is still unclear in mice (Ober et al. 2004; Matsui et al. 2005; Zerbe et al. 2008). If cells are unable to migrate out of the node, they will not become endoderm. If cells migrate at inappropriate times, regions are not specified correctly. Cell position controls exposure of cells to signaling molecules from the surrounding tissue. Several transcription factors are critical for cell movement out of the primitive streak and thus the formation of regional identity. The mechanisms of early regionalization are not fully defined; however, several factors have been shown to be critical.
Initial specification of the definitive endoderm and morphogenesis requires the transcription factor, Sox17 (a SRY-related HMG factor), in multiple species (Hudson et al. 1997; Alexander and Stainier 1999; Clements and Woodland 2000; Kanai-Azuma et al. 2002). Sox17 expression is high in all definitive endoderm cells early on. Sox17 was shown to cooperate with Wnt signaling, and to activate Foxa2 (a member of the forkhead transcription factor family) (Sinner et al. 2004). Subsequently expression of Sox17 is restricted to the posterior end of the embryo and Sox17 null cells are incapable of forming mid and hindgut (Kanai-Azuma et al. 2002). Even later Sox17 interacts with another transcription factor, Pdx1 (pancreatic and duodenal homeobox 1), which is required for the specification of the pancreas (Spence et al. 2009). Using both loss of function and gain of function techniques in the mouse embryo, Sox17 was shown to repress Pdx1 expression in the liver primordium, a process that is critical for establishing organ domain boundaries between liver and pancreas (Spence et al. 2009).
The anterior endodermal region requires two major transcription factors, FoxA2 and Hhex. FoxA2 is the master regulator of the anterior primitive gut. FoxA2 mutant mice show defects in cell migration after endodermal specification and thus loss of all foregut and midgut structures; however, hindgut development is unaffected (Weinstein et al. 1994; Dufort et al. 1998). Using tetraploid embryo complementation, it was later shown that Foxa2 null cells can form the hindgut but cannot incorporate into the developing fore- or midgut (Dufort et al. 1998). Hhex expression is required for anterior endoderm development and activated by both Nodal and Wnt signaling (Martinez-Barbera et al. 2000; Smithers and Jones 2002). The promoter of Hhex has been shown to have both activation and repression domains that are responsive to multiple signaling pathways including Nodal, Wnt and BMP (Rodriguez et al. 2001; Rankin et al. 2011). Sox2 is also required in a dose-dependent manner in the developing foregut (Que et al. 2007). All of these factors are important throughout the morphogenesis of the anterior part of the gut and will be discussed in more detail.
The Caudal-related homeobox transcripton factor Cdx2 is required for posterior gut development. Cdx2 expression is highest at E8.5 in the hindgut, and its expression in the intestinal epithelium defines a clear-cut boundary at the foregut-midgut junction. Cdx2 is absolutely required in the midgut and hindgut for the formation of the intestine (Gao et al. 2009). Studies in Caco-2 cells, a colon cancer cell line that is used as a model for the transition of cells from progenitor to differentiation in the intestine, have shown the Cdx2 regulates expression of both progenitor and differentiation specific genes (Verzi et al. 2010a) (Gao et al. 2009). Gene regulation may be due to a role of Cdx2 in chromatin accessibility at these loci (Verzi et al. 2010b). Cdx2 is also required for homeostasis of mature intestinal epithelial cells, which will be discussed in detail in the following sections.
2. Signaling across tissues modulates regional transcription factor activity
Signaling from the mesoderm maintains hindgut fates and actively represses foregut development in the posterior endoderm, at least in Xenopus development (Zorn and Wells 2007). Wnt signaling, well known for its role in establishing the anterior-posterior axis of the embryo (Huelsken et al. 2000), is highly active in the hindgut and represses foregut identity (McLin et al. 2007). Similarly, bone morphogenetic protein (BMP) signaling is required for hindgut development, and naturally occurring antagonists are required to allow foregut development (Sasai et al. 1996; Zorn et al. 1999; Tiso et al. 2002). In addition, BMP signaling has been shown to be important in determining cell fates in the foregut during organogenesis, which will be discussed in more detail below. Fibroblast growth factor (FGF) seems to be expressed in a gradient, with highest expression in the posterior gut, and it represses anterior markers (Serls et al. 2005; Dessimoz et al. 2006). However, varying concentrations of FGF are also required for different lineages that arise from the ventral foregut (Jung et al. 1999; Calmont et al. 2006; Zaret and Grompe 2008).
Retinoic acid signaling has multiple roles in establishing anterior-posterior regional identity. Studies in mice deficient for retinoic acid signaling in the foregut through deletion of the retinaldehyde dehydrogenase 2 (Raldh2) gene or treatment with a pan-retinoic acid receptor (RAR) antagonist, show failure to develop multiple anterior organs (Wendling et al. 2000; Molotkov et al. 2005; Wang et al. 2006). Retinoic acid seems to act through regulation of transcription factors with retinoic acid responsive enhancers including Hoxb1 and Hoxa5 (Huang et al. 1998; Matt et al. 2003; Niederreither et al. 2003; Grapin-Botton 2005). However, there may be multiple additional effects of retinoic acid signaling including activation of other signaling factors such as FGF10 and Sonic hedgehog (Shh) (Ivins et al. 2005; Wang et al. 2006).
3. Combinations of transcription factors determine organ domains in the primitive gut tube
Extensive investigation of the expression of transcription factors has been performed in an effort to understand the establishment of the multiple organ domains in the gastrointestinal system. For example, a study of 15 transcription factors expressed within the developing mouse foregut identified over a dozen unique domains that roughly correspond with particular organs (Sherwood et al. 2009). The mechanism behind the complex combinatorial control is organ specific and will be discussed in the following sections.
1. Development of the Intestine
As the foregut organs are being specified, the gut tube is further regionalized by epithelial-mesenchymal interactions that establish gene expression throughout the gastrointestinal tract. The small and large intestines are specified from the midgut and hindgut, respectively. Even though small and large intestine form a continuous tube and share similar developmental origins, their morphology and final functions are unique. Close association with the mesoderm is required for both development and adult function.
The adult intestinal epithelium is one of several epithelial tissues in the body that maintains its function by constant production of several types of short-lived cells. The intestinal epithelium is composed of a single layer of cells that perform the essential role of digestion and absorption of nutrients into the blood stream. Epithelial cells migrate from the crypt region to the villus, changing from a progenitor state to fully differentiated cell in the process. Once cells reach the tip of the villus, they are shed into the gut lumen. This entire process takes approximately 3–5 days in mice and human. Four major differentiated cell types are required to maintain small intestinal function. 90% of all epithelial cells are enterocytes that function as absorptive cells. The remainder of the cells are composed of enteroendocrine, goblet and Paneth cells, collectively referred to as the secretory cell lineage. These cells secrete hormones that regulate digestion and signal to the body, elaborate mucous that protects the epithelium, and produce defensins to protect against infections, respectively. The mature epithelial cell types must be maintained in the appropriate ratio or there are severe consequences for intestinal function. Interestingly, the relative representation of the four differentiated cell types also varies across the anterior-posterior axis, with the duodenum, the most anterior section of the intestine, elaborating far fewer goblet cells than the colon, while Paneth cells are found in the small intestine but are missing from the large bowel. Thus, positional cues must maintain differences in progenitor cell differentiation even in the adult.
2. Regional specification and morphogenesis of the intestine
Cdx2 is one of the earliest transcription factors expressed in the primitive gut tube and is required for defining both midgut and hindgut regions that contribute to the entire intestine. Cdx2 is expressed most highly in the hindgut but its expression extends all the way to the foregut-midgut boundary (Silberg et al. 2000). In fact, while the very first duodenal epithelial cell is Cdx2-positive, all cells of the stomach and esophagus lack Cdx2. For this reason, Cdx2 expression is frequently employed as a marker of intestinal metaplasia, a precursor to cancer. Specification of the colon and expression of many intestine-specific genes require Cdx2 (Gao et al. 2009; Gao and Kaestner 2010) (Verzi et al. 2010a; Verzi et al. 2010b). Regulation of the Hox gene expression along the A-P axis further specifies the intestine and defines areas of major anatomical constrictions (Kawazoe et al. 2002; Grapin-Botton 2005; Hanamura et al. 2006). Cdx2 also impacts the Hox code (Gao et al. 2009).
Multiple signaling pathways converge on Cdx2 to regulate intestinal development. Wnt persistent in the hindgut regulates the expression of Cdx2. Mutations in Wnt signaling cause lower expression of Cdx2 and show an increased area of established foregut (Gregorieff et al. 2004; Cervantes et al. 2009). FGF signaling also plays a role in the establishment of the Cdx boundary at the duodenal-pyloric junction (Dessimoz et al. 2006; Rubin 2007; Benahmed et al. 2008).
Hedgehog signaling is also important for interactions between the mesenchyme and endoderm for intestinal specification and regionalization. Mutants in both Sonic hedgehog (Shh) and Indian hedgehog (Ihh) show defects in the gastrointestinal tract, including intestinal transformation of the stomach (Ramalho-Santos et al. 2000; van den Brink 2007; Saqui-Salces and Merchant 2010). In addition, ectopic expression of Shh causes transformation of pancreas into intestine (Apelqvist et al. 1997).
The simple endodermal pseudo-stratified epithelium of the primitive gut begins to transform into the mature intestinal columnar epithelium beginning at day 14 of gestation in mice, a process termed the epithelial transition (Figure 2). Villi are projections into the lumen that are morphologically different but contiguous with the crypts, which are invaginations into the supporting mesenchyme. Villi are formed from polarized folds of endoderm while highly proliferative pockets of cells form between folds that are the beginnings of crypt formation (Abud and Heath 2004; Abud et al. 2005). These morphological changes require tight interactions between the epithelium and underlying mesenchyme, as seen, for instance, by a delay in epithelialization in the absence of the mesenchymal transcription factor FoxL1 (Kaestner et al. 1997). Communication between these tissues requires Hh, Wnt, and BMP signaling (Li et al. 2007; Madison et al. 2009).
Figure 2
Figure 2
Intestinal morphology changes during development.
3. Homeostasis in the intestinal epithelium
The intestinal epithelium maintains its self-renewal capacity by maintaining a multipotent stem cell niche. The intestinal epithelium is constantly repopulated by the coordinated division of stem cells into faster cycling transit amplifying cells that divide to produce all differentiated cells (Sancho et al. 2004). Intestinal stem cells are found in the bottom of the crypts and divide symmetrically to produce both stem cells and transit amplifying cells (Snippert et al. 2010). These cells are bona fide stem cells and are sufficient to form new crypts in culture (Sato et al. 2009). Intestinal stem cells also express general stem cell markers such as Lgr5 (Barker et al. 2010), bmi-1 (Sangiorgi and Capecchi 2008), prominin/CD133 (Zhu et al. 2009; Snippert et al. 2010), and DCMKL-1 (May et al. 2009); however, little is known how or if these markers themselves contribute to maintenance of self-renewal. However, the Lgr5 relative, Lgr4, is expressed in the crypt epithelium as well as the surrounding mesenchyme and was shown using ex vivo culture techniques to be required within the epithelium for maintenance of the crypts (Mustata et al. 2011).
Wnt signaling is required for maintenance of undifferentiated cells. There is a gradient of Wnt expression with highest levels found at the bottom of the crypt, which gradually decreases as cells transit up (van de Wetering et al. 2002; Pinto et al. 2003; Ireland et al. 2004; Sansom et al. 2004). Many Wnt responsive genes also show highest activation in the crypt region (Van der Flier et al. 2007). Mutations that cause excessive activation of Wnt signaling in the intestinal epithelium, such as those found in the adenomatos polyposis coli (APC) gene, cause massive growth in the epithelium and proliferation into cancerous polyps (Fearon 2011). In fact, greater than 80% of human colorectal cancers show mutations in the APC gene or other genes in the Wnt/APC/beta-catenin pathway.
Multiple signaling pathways are critical regulators of differentiation. Active Notch signaling and BMP promote differentiation into specific intestinal cell types. Underlying the epithelium is mesenchymal tissue that serves as both structural support and signaling center. Expression of Hedgehog and BMP ligands serve as a way for mesenchyme and epithelium to communicate through reciprocal signaling, and disruption of either causes defects in proliferation (Crosnier et al. 2006; Madison et al. 2009). These interactions between cells are important because support cells such as myofibroblasts enhance survival and growth of intestinal epithelium in vitro culture (Ootani et al. 2009). It is unclear how signaling pathways interact to regulate gene expression as cells transition from stem to differentiated states within the epithelium, because the concentrations of the various ligands have to be modulated across the very small distances that separate stem, progenitor, and differentiated cells.
Positioning within the crypt/villus axis and cell migration are essential for regulation of proliferation and differentiation. EphB and Ephrin-B levels vary with position along the crypt/villus axis (Batlle et al. 2002). The migratory behavior of the cell is tightly correlated with its differentiation status (Wimmer-Kleikamp et al. 2004; Pasquale 2005; Vearing and Lackmann 2005). Components of the Eph-ephrin signaling pathway are targets of Wnt/β-catenin signaling (Batlle et al. 2002). Compound EphB2/EphB3 mutant mice show differentiated cells occupying positions in the proliferative zone and a reduced proliferative zone, suggesting that the EphB receptors play a role in repelling the downward migration of differentiated cells (Batlle et al. 2002). Recent evidence has shown that Eph-ephrin signaling is also dependent on other signaling pathways (Genander et al. 2009; Koo et al. 2009; Furukawa et al. 2011).
As mentioned above, intestinal epithelial cells are classified as either absorptive enterocytes (termed colonocytes in the colon) or secretory cells (goblet, Paneth and enteroendocrine cells). Notch signaling activates Hes1 that antagonizes enterocyte fate. Notch works by lateral inhibition to prevent adjacent cells from adopting the same fate as the signal-emitting cell thus controlling the final composition of differentiated cell types (Artavanis-Tsakonas et al. 1999; Gaiano et al. 2000). Secretory cells depend on Math1 expression in progenitor cells (Yang et al. 2001; Shroyer et al. 2007; VanDussen and Samuelson 2010). Math1 then activates the expression of Sox9, Klf4 and NeuroD/Ngn3 to direct full differentiation into Paneth, goblet and enteroendocrine cells, respectively (Naya et al. 1997; Jenny et al. 2002; Katz et al. 2002; Lee et al. 2002; Mori-Akiyama et al. 2007). More detail on intestinal epithelial differentiation can be found in a recent review (May and Kaestner 2010).
1. Development of the organ primordia
After regional patterning is established, organ domains are specified. The foregut domain gives rise to many of the major organs in the gastrointestinal tract, including lungs, stomach, pancreas and liver. Complex patterns of transcription factors are associated with each organ domain. Activation of transcription factors is due in part to signaling from adjacent tissues. However, the mechanism of how these combinations of signals cause gene expression and morphology changes into organ primordia is still unclear. The process of the development of the organ primordia has been reviewed elsewhere in detail (Zorn and Wells 2009).
The establishment of the final morphology of the gut-associated organs follows both similar and diverse programs. All gut-associated organs - also called the para-alimentary tract - use signals from adjacent tissues to invade the local mesenchyme adjacent to the primitive gut tube to form an organ bud. At this point, all gut-associated organs follow unique programs that allow for proliferation and differentiation. The lung depends on endoderm-mesenchymal interactions to direct its branching structure and generation of several functional cell types, with major contributions from FGF and TGF-β signaling (Maeda et al. 2007). The pancreas forms polarized microlumina that eventually coalesce to form the final ductal tree, a process that is totally independent of mesenchymal interactions (Gittes 2009; Villasenor et al. 2010). The liver has a close association with the vasculature and generates bi-potential progenitors that differentiate into a homogeneous population of functional cells (Zaret and Grompe 2008; Nagaoka and Duncan 2010). Here we focus on liver organ formation as an example due to the large amount of information that is known about the transcriptional regulation of liver development.
2. Setting up transcription factor networks in the hepatic primordium
As mentioned previously, transcription factors are required not only for initial specification of the endoderm at gastrulation, but are continually involved throughout liver development. FoxA1 and FoxA2, act in concert to enable the induction of the hepatic gene program (Lee et al. 2005). These transcription factors are thought to function as pioneer factors by facilitating the opening of chromatin at several important liver-specific genes, including albumin and alpha-fetoprotein (Gualdi et al. 1996; Zaret 1996; Cirillo et al. 1998; Crowe et al. 1999). The GATA family of zinc finger transcription factors, GATA 4 and 6, also act together in hepatic gene induction, including the activation of the albumin locus (Bossard and Zaret 1998; Cirillo et al. 1998), and subsequent liver development requires the presence of at least one of them (Holtzinger and Evans 2005; Zhao et al. 2005).
Hepatic fate is controlled in addition by signals from the surrounding mesenchyme. FGF signaling from the cardiac mesoderm activates MAPK signaling that induces hepatic gene induction (Rossi et al. 2001; Chen et al. 2003; Zhang et al. 2004; Serls et al. 2005; Calmont et al. 2006; Shin et al. 2007). BMP 2 and 4 signals from the septum transversum mesoderm enhance the hepatic competence of the endoderm (Jones et al. 1991; Smith and Harland 1992; Furuta et al. 1997). TGF-β acts as a developmental timer to maintain hepatocyte competency while restricting differentiation until endodermal cells are positioned correctly (Wandzioch and Zaret 2009). Wnt signaling is required for liver bud development and hepatic growth through the activation of transcription factors such as Hhex (Finley et al. 2003; Monga et al. 2003; Suksaweang et al. 2004).
3. Tissue patterning and morphogenesis
Liver bud formation takes place on the ventral wall of the foregut endoderm and requires several morphological changes. Cells positioned near the developing heart receive signals that are critical for epithelial thickening and formation of the liver bud outgrowth from the endoderm (Douarin 1975). This signal is mediated, at least in part, by FGF, as demonstrated through in vitro culture studies (Gualdi et al. 1996). These endodermal cells then delaminate and invade the septum transversum mesenchyme (STM) to begin the formation of the final organ. Hepatoblasts subsequently fully differentiate into functional cell types that are discussed below.
Cell migration is dependent on two homeobox transcription factors, HHex and Prox1. Hhex is a transcriptional repressor and required for hepatoblast proliferation, but not their initial specification (Keng et al. 2000; Martinez Barbera et al. 2000). Hhex is also required for liver induction by positioning the ventral endoderm within the cardiogenic field (Martinez-Barbera et al. 2000; Bort et al. 2004; Hunter et al. 2007). Interestingly, loss of Prox1, a prospero-related homeobox transcription factor, has no effect on liver-specific gene expression; however, the levels of the cell adhesion molecule, E-cadherin, are increased dramatically in Prox1 mutant embryos, preventing hepatoblasts to delaminate from the ventral foregut (Sosa-Pineda et al. 2000). Thus, it is apparent that transient migratory behavior is required for liver development.
4. Diversification and homeostasis in the liver
Hepatoblasts are the earliest differentiated liver cell type. These cells function as progenitors and are able to differentiate fully two distinct functional cell types. Expression of Albumin (Alb), transthyretin (Ttr) and alpha-fetoprotein (Afp) are the earliest markers of hepatoblasts (Gualdi et al. 1996; Jung et al. 1999). Hepatoblasts are bipotential cells that further differentiate into mature epithelial cell types, the hepatocytes that form the main functional cell of the liver and the cholangiocytes that form the biliary tree.
Cholangiocytes are cells that line the bile ducts and function in synthesizing and secreting components of bile and make up a small percentage of the liver. Hnf6, hepatic nuclear factor 6, is required for the formation of biliary ducts (Clotman et al. 2002). In the liver, Hnf6 transactivates the promoter of another transcription factor, Hnf1b, which is required generally for the development of tubules during organogenesis (Clotman et al. 2002; Coffinier et al. 2002). The exact mechanism of this transcriptional cascade is still being studied but most likely is regulated by signals from the septum transversum mesenchyme (Kalinichenko et al. 2002).
Hepatocytes make up approximately 78% of the total liver volume (Blouin et al. 1977). They are a polarized epithelial cell type that have many functions including controlling the levels of metabolites and serum proteins in the blood (Stamatoglou and Hughes 1994). HNF-4 is critical for terminal hepatocyte differentiation, although is not required for early liver specification (Li et al. 2000). HNF-4 may activate transcription of targets directly through regulation of chromatin accessibility (Li et al. 2000; Soutoglou and Talianidis 2002). HNF-4 also targets genes indirectly through the activation of the transcriptional regulators Hnf1a and PXR, which are crucial for expression of subsets of hepatocyte specific genes (Tian and Schibler 1991; Kuo et al. 1992; Holewa et al. 1996). HNF-4 is also required for the epithelialization of the liver (Spath and Weiss 1998; Battle et al. 2006; Hayhurst et al. 2008).
A complex transcriptional network is required for liver development. These factors include HNF-1a, HNF-1B, FoxA1, FoxA2, FoxA3, HNF-4, COUP-TFII, LRH-1, FXRa, PXR, C/EBPa and HNF-6 (Cereghini 1996; Costa et al. 2003). Multiple transcription factors bind to hepatic gene promoters to induce robust expression (Figure 3A). For instance, promoters of active hepatic genes that were bound with HNF-1 or HNF-6 were also often occupied by HNF-4a (Odom et al. 2004). These factors also have reciprocal regulation in which expression of one factor depends on another factor in the same cell (Figure 3B) (Kuo et al. 1992; Bulla 1997; Bailly et al. 1998). Investigation of the promoter occupancy and expression patterns of these transcription factors during liver development revealed that the increased number of interactions is correlated with hepatocyte differentiation and increased expression of individual transcription factors within the network (Kyrmizi et al. 2006).
Figure 3
Figure 3
Regulation of liver-specific gene expression using a transcriptional network
Much work has been done to investigate the ability of the adult liver to regenerate, as this process is important in liver transplantation. The liver can fully regenerate after an acute injury, for instance the surgical removal of 70% of the liver mass (partial hepatectomy). However, the ability for the liver to recover after chronic injury is often compromised. Multiple signaling pathways including FGF, BMP, Wnt, and Notch have been shown to be involved in this recovery process of the liver (Bohm et al. 2010). This rapid growth process is largely due to hepatocyte replication (Evarts et al. 1987; Evarts et al. 1989). Much work has focused in hemapoietic stem cells being the source of regenerative capacity in the liver; however, these studies have mixed results (Schwartz and Verfaillie 2010). Recently the search for an adult facultative liver progenitor cell has moved forward with the discovery of a very small population of Foxl1-positive cells that expand during liver injury and are able to differentiate into all liver cell types after injury (Sackett et al. 2009; KHK, pers. comm.). One can envision that in the future the isolation and ex vivo expansion and differentiation of these hepatic progenitors might be put to therapeutic use.
Concluding Remarks
This chapter highlights the complex development of the gastrointestinal tract. Regionalization of the primitive gut provides a framework in which organ domains are established. Interactions between the endoderm and mesoderm provide transcriptional activation of diverse genes across regions to form a mature intestine. Combinatorial control of gene expression by complex transcriptional networks are required for liver development and homeostasis.
Acknowledgments
Work in the Kaestner lab related to the findings described above have been supported by NIH grants DK049210 and DK053839.
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