Glucose Transporter Expression in an Avian Nectarivore: The Ruby-Throated Hummingbird (Archilochus colubris)

Abstract

Glucose transporter (GLUT) proteins play a key role in the transport of monosaccharides across cellular membranes, and thus, blood sugar regulation and tissue metabolism. Patterns of GLUT expression, including the insulin-responsive GLUT4, have been well characterized in mammals. However, relatively little is known about patterns of GLUT expression in birds with existing data limited to the granivorous or herbivorous chicken, duck and sparrow. The smallest avian taxa, hummingbirds, exhibit some of the highest fasted and fed blood glucose levels and display an unusual ability to switch rapidly and completely between endogenous fat and exogenous sugar to fuel energetically expensive hovering flight. Despite this, nothing is known about the GLUT transporters that enable observed rapid rates of carbohydrate flux. We examined GLUT (GLUT1, 2, 3, & 4) expression in pectoralis, leg muscle, heart, liver, kidney, intestine and brain from both zebra finches (Taeniopygia guttata) and ruby-throated hummingbirds (Archilochus colubris). mRNA expression of all four transporters was probed using reverse-transcription PCR (RT-PCR). In addition, GLUT1 and 4 protein expression were assayed by western blot and immunostaining. Patterns of RNA and protein expression of GLUT1-3 in both species agree closely with published reports from other birds and mammals. As in other birds, and unlike in mammals, we did not detect GLUT4. A lack of GLUT4 correlates with hyperglycemia and an uncoupling of exercise intensity and relative oxidation of carbohydrates in hummingbirds. The function of GLUTs present in hummingbird muscle tissue (e.g. GLUT1 and 3) remain undescribed. Thus, further work is necessary to determine if high capillary density, and thus surface area across which cellular-mediated transport of sugars into active tissues (e.g. muscle) occurs, rather than taxon-specific differences in GLUT density or kinetics, can account for observed rapid rates of sugar flux into these tissues.

Glucose Transporter Expression and Regulation Following a Fast in the Ruby-throated Hummingbird,Archilochus colubris

“…In this study, 143 using custom antibodies for the different isoforms of hummingbird GLUTs, we sought to 144 identify the tissue-specific protein distribution and to quantify the abundance in the PM, of 145 GLUT1, GLUT2, GLUT3, and GLUT5. We predicted GLUT1 would be detected in 146 hummingbird flight muscle, cardiac, and liver tissue, in accordance with its ubiquitous presence 147 in mammalian tissue (Mueckler and Thorens, 2013), as well as its previous detection in 148 hummingbird myocytes (Welch et al, 2013). As GLUT2 plays a stronger role in enteric 149 (Karasov, 2017) and hepatic (Mueckler and Thorens, 2013) sugar transport, we predicted that its 150 abundance would be limited in muscles and more predominantly found in the liver.…”

“…GLUT1 protein in WT 531 liver homogenates of ruby-throated hummingbirds was only minimally visible (Table 1) and 532 was, surprisingly, not detected in PM fractions (Table 2). This result is in contrast to previously 533 reported detection of hepatic mRNA transcript for GLUT1 in both chickens (Byers et al, 2018) 534 and hummingbirds (Welch et al, 2013). However, as GLUT1 is abundant in erythrocytes 535 (Carruthers, 2009), it is possible that the previous mRNA detection, as well as our detection of 536 some hepatic GLUT1 protein, may have resulted from red blood cell contamination.…”

“…In mammals, GLUT4 translocation to the PM by 127 insulin-stimulation following feeding is known to recruit other GLUT isoforms to the PM as 128 well, increasing the sugar import rate into active cells (Guma et al, 1995). Hummingbirds 129 (Welch et al, 2013), much like chickens (Byers et al, 2018), do not express transcript or protein 130 of the insulin-sensitive GLUT4 isoform. Chicken insulin levels do not significantly change with 131 though it may in the liver (Dupont, 2009;Zhang et al, 2013).…”

“…Previous studies have confirmed the presence of GLUT1 and GLUT5 transcript in nearly 140 all hummingbird tissue examined . Immunohistochemistry of 141 hummingbird myocytes using a commercial antibody for GLUT1 have also shown GLUT1 142 localisation to the PM (Welch et al, 2013), though, the results were not definitive. In this study, 143 using custom antibodies for the different isoforms of hummingbird GLUTs, we sought to 144 identify the tissue-specific protein distribution and to quantify the abundance in the PM, of 145 GLUT1, GLUT2, GLUT3, and GLUT5.…”

Single molecule, full-length transcript sequencing provides insight into the extreme metabolism of ruby-throated hummingbirdArchilochus colubris

“…The UCP homolog was cloned from pectoralis (flight muscle) and its functional significance in vivo is unclear. The amino acid sequence of hummingbird insulin was found to be largely identical to that from chicken; however, birds are insulin insensitive and lack the insulin-regulated glucose transporter (GLUT) protein GLUT4, making the role of this hormone in the regulation of energy homeostasis in hummingbirds unknown (Welch et al 2013;Braun and Sweazea 2008;Polakof et al 2011).…”