© 2001 American Heart Association, Inc.
Editorials |
Road Rage
Cardiac Rab1 and ER-to-Golgi Traffic
From the Center for Cardiovascular Research, Departments of Medicine, Cell Biology & Physiology, Washington University School of Medicine, St. Louis, Mo.
Correspondence to Anthony J. Muslin, MD, Center for Cardiovascular Research–Box 8086, 660 South Euclid Ave, Saint Louis, MO 63110. E-mail amuslin{at}im.wustl.edu
Key Words: Rab • Golgi • endoplasmic reticulum • transgenic • congestive heart failure
Ras-related low molecular weight guanosine triphosphatases (GTPases) regulate nearly all aspects of cell physiology including growth, differentiation, proliferation, cell movement, and nuclear transport. The Rab family of GTPases regulates another aspect of physiology: the transport of membranes throughout the cell.1–5 There are at least 30 members of the Rab family in mammalian species, and different Rabs regulate different aspects of tubulovesicular trafficking.1,2 For example, Rab1 regulates the transport of newly synthesized proteins from the rough endoplasmic reticulum (ER) to the Golgi in the exocytic pathway1–5 (Figure). Most secreted proteins and transmembrane receptors pass through this exocytic pathway. Rough ER-to-Golgi transport initially involves the formation of vesicles from the ER under the direction of a protein coat called COPII.5 The COPII coat bends the ER membrane so that it pinches off as a vesicle (Figure). The vesicle quickly loses its protein coat and then moves along microtubule tracks to the cis-Golgi complex where the vesicle fuses and delivers its protein cargo.4 Secretory and membrane proteins then sequentially travel through the cis, medial, and trans compartments of the Golgi complex during which the proteins undergo posttranslational modifications before their transit to the extracellular space or to the plasma membrane.
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Rab GTPases play a key role in tubulovesicular trafficking by regulating key membrane targeting and fusion events.1–5 Rab proteins are lipid modified, and this usually consists of double geranylgeranylation near their carboxy termini, which allows them to bind avidly to lipid membranes.1,2 Specific guanine nucleotide exchange factors activate Rabs, by replacing GTP for GDP. Once activated, Rab-GTP regulates vesicular fusion events by coordinating the linkage of cognate v-SNAREs (soluble N-ethylmaleimide–sensitive factor attachment protein receptors of the VAMP family) with t-SNAREs (syntaxin family), perhaps via tethering factors such as p115.5,6 SNARE proteins are assembled into a complex that is typically composed of four 
helices contributed by up to four distinct SNAREs, with all of the transmembrane domains localized at one end of the bundle.7 Formation of a SNARE complex that links a vesicle to its target membrane results in membrane fusion. After membrane fusion, the GTP on Rab is hydrolyzed to GDP, and inactive Rab-GDP is bound by a Rab GDP-dissociation inhibitor (RabGDI) that recycles the Rab back to its membrane of origin.1,2
The prototypical Rab protein is Ypt1p, a budding yeast protein that is essential for survival and that regulates rough ER-to-Golgi transport by facilitating SNARE complex formation. Interestingly, overexpression of several SNARE component genes can compensate for the loss of Ypt1p.8,9 It is possible that overexpression of a SNARE might abrogate the need for a Rab that facilitates SNARE pairing by mass action. Mammalian Rab1 performs a similar function in the regulation of rough ER-to-Golgi transport in the exocytic pathway. Distinct Rabs regulate other membrane fusion events. For example, Rab6 regulates retrograde vesicular transport from the Golgi apparatus to the ER, and Rab4 regulates plasma membrane recycling of proteins.1–5
Virtually nothing is known about the role of Rab proteins in cardiac cell biology. To address this knowledge gap, Wu and coworkers10 investigated the expression and function of Rab proteins in murine cardiac tissue in the present issue of Circulation Research. They found that Rabs 1b, 3, 4, 5, and 6 were highly expressed in adult murine and human cardiac tissue. In addition, Rab1a was expressed in human cardiac tissue. To determine whether altered expression of Rab genes correlated with cardiac dysfunction, Wu et al examined transgenic mice with cardiac-specific overexpression of the ß2 adrenergic receptor (AR). In transgenic mice with 60-fold overexpression of the ß2AR, basal cardiac function is enhanced without an increase in mortality rate at 1 year of age.11 However, transgenic mice with 350-fold overexpression of ß2AR develop a rapidly progressive form of dilated cardiomyopathy and die at 25 weeks of age. In the high-level ß2AR-overexpressing line, expression of Rabs 1b, 4, and 6 was significantly increased. This increase in Rab expression may occur because the large amount of cardiac ß2AR necessitates enhanced protein trafficking machinery.
To determine whether the source of cardiac dysfunction in ß2AR transgenic mice was the increased expression of Rab genes, and not a direct result of increased adrenergic signaling, Wu et al10 generated transgenic mice with cardiac-specific overexpression of Rab1a. Their choice of Rab1a was governed by the fact that Rab1, as the functional and structural homologue of Ypt1p, is the best studied of all of the Rabs. Overexpression of Rab1a did not alter the expression of other Rabs in the heart, nor did it affect the level of expression of two cardiac RabGDIs. Increased expression of wild-type Rab1a in cardiac tissue resulted in a profound and reproducible phenotype in high expressors that consisted of a dilated cardiomyopathy that resulted in premature death at 
6 weeks of age. Hearts of high-expressor Rab1a transgenic mice at 6 weeks were dilated with prominent atrial thrombi and also showed evidence of cardiac hypertrophy. In addition, the expression of atrial natriuretic factor (ANF), ßMHC, and -skeletal actin was increased in high-expressor transgenic heart tissue at 6 weeks of age. Furthermore, there was altered expression of three protein kinase C (PKC) isoforms, PKC, 
, and 
, in medium-expressor cardiac tissue, and an increase in the particulate fraction localization of PKC
. Despite the gross enlargement of the cardiac chambers that was observed in the high-expressor line, there was no evidence of cardiac myocyte apoptosis by TUNEL staining.
Ultrastructural analysis of ventricular myocytes from Rab1a medium expressors before the onset of functional deterioration revealed that Golgi stacks and surrounding transitional vesicles were markedly enlarged. Atrial myocytes from high-expressor transgenic mice were notable for a large increase in the number of vesicles containing ANF granules. Immunogold labeling of ultrathin sections revealed that Rab1a was associated with the abnormal vesicular structures observed in ventricular myocytes. One explanation for the abnormal vesicular structures observed in transgenic myocytes is that isolated overexpression of Rab1a results in the depletion of tethering factors or SNAREs so that vesicular fusion to and within the Golgi complex becomes inefficient. Despite these profound ultrastructural alterations observed in Rab1a transgenic mice, it is not immediately apparent whether the abnormal vesicular structures observed or the increased expression of PKC isoforms is causally linked to cardiac dysfunction.
The work of Wu et al10 establishes that several Rab family members are expressed in mammalian cardiac tissue and that marked overexpression of a transmembrane receptor, the ß2AR, can result in the altered expression of several Rab family members. Their work also shows that overexpression of Rab1a in murine cardiac tissue is sufficient to cause profound cardiac dysfunction with chamber enlargement, cardiac hypertrophy, and premature death. It is not clear whether increased Rab1 expression is required for the ß2AR to cause cardiac dysfunction in mice. Nor is it clear that altered Rab1 expression or activity is associated with—or causally linked to—human cardiac disease. Nevertheless, this work represents an important first step in the elucidation of the role of Rab proteins in cardiac function and disease.
Footnotes
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.
References
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Wu G, Yussman MG, Barrett TJ, Hahn HS, Osinska H, Hilliard GM, Wang X, Toyokawa T, Yatani A, Lynch RA, Robbins J, Dorn GWII. Increased myocardial Rab GTPase expression: a consequence and cause of cardiomyopathy. Circ Res. 2001; 89: 1130–1137.
11.
Liggett SB, Tepe NM, Lorenz JN, Canning AM, Jantz TD, Mitarai S, Yatani A, Dorn GW2nd. Early and delayed consequences of ß2-adrenergic receptor overexpression in mouse hearts: critical role for expression level. Circulation. 2000; 101: 1707–1714.
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