Aestivation Induces Changes in the mRNA Expression Levels and Protein Abundance of Two Isoforms of Urea Transporters in the Gills of the African Lungfish, Protopterus annectens

Abstract

The African lungfish, Protopterus annectens, is ammonotelic in water despite being ureogenic. When it aestivates in mucus cocoon on land, ammonia is detoxified to urea. During the maintenance phase of aestivation, urea accumulates in the body, which is subsequently excreted upon arousal. Urea excretion involves urea transporters (UT/Ut). This study aimed to clone and sequence the ut isoforms from the gills of P. annectens, and to test the hypothesis that the mRNA and/or protein expression levels of ut/Ut isoforms could vary in the gills of P. annectens during the induction, maintenance, and arousal phases of aestivation. Two isoforms of ut, ut-a2a and ut-a2b, were obtained from the gills of P. annectens. ut-a2a consisted of 1227 bp and coded for 408 amino acids with an estimated molecular mass of 44.7 kDa, while ut-a2b consisted of 1392 bp and coded for 464 amino acids with an estimated molecular mass of 51.2 kDa. Ut-a2a and Ut-a2b of P. annectens had a closer phylogenetic relationship with Ut/UT of tetrapods than Ut of fishes. While the mRNA expression pattern of ut-a2a and ut-a2b across various tissues of P. annectens differed, the transcript levels of ut-a2a and ut-a2b in the gills were comparable, indicating that they might be equally important for branchial urea excretion during the initial arousal phase of aestivation. During the maintenance phase of aestivation, the transcript level of ut-a2a increased significantly, but the protein abundance of Ut-a2a remained unchanged in the gills of P. annectens. This could be an adaptive feature to prepare for an increase in the production of Ut-a2a upon arousal. Indeed, arousal led to a significant increase in the branchial Ut-a2a protein abundance. Although the transcript level of ut-a2b remained unchanged, there were significant increases in the protein abundance of Ut-a2b in the gills of P. annectens throughout the three phases of aestivation. The increase in the protein abundance of Ut-a2b during the maintenance phase could also be an adaptive feature to prepare for efficient urea excretion when water becomes available.

Effects of Experimental Terrestrialization on the Skin Mucus Proteome of African Lungfish (Protopterus dolloi)

“…As the water dissipates completely, the mucus/mud mixture coating around the lungfish body hardens forming a cocoon that protects the animal for months. Once encased, the lungfish ceases feeding and locomotive activities, has to prevent cell death, and sustain a slow rate of waste production until conditions become favorable ( 11 ), this is the maintenance phase of aestivation, where the lungfish can lay dormant for years. Upon the introduction of water, the lungfish instantly awakens from dormancy, leaves the mucus cocoon, and slowly swims toward the surface of the water for air ( 5 ).…”

Molecular characterization of three Rhesus glycoproteins from the gills of the African lungfish, Protopterus annectens, and effects of aestivation on their mRNA expression levels and protein abundance

“…annectens , our results are in agreement with the notion that lungfish is the intermediary form between fish and tetrapods. Since lungfishes are the closest living sister group of land vertebrates, they would logically possess some genes/proteins that are closer to those of other fishes (Cps III, [ 11 ]; Ass, [ 12 ]) and others that have greater similarity to those of tetrapods (Asl, [ 12 ]; L-gulono-γ-lactone oxidase, [ 30 ]; Na + /K + -ATPase α-subunit isoforms, [ 31 ]; Coagulation factor II and Fibrinogen gamma chain [ 32 ]; Betaine-homocysteine S -methyltransferase 1, [ 33 ]; Myostatin [ 34 ]; Aquaporin 1 and 3 [ 24 ]; Urea transporter isoforms [ 29 ]).…”

“…The protein loads of the gill samples for gel separation were 100 μg for Rhag, 100 μg for Rhbg and 50 μg for Rhcg. Aestivation involves a complex interplay between up-regulation and down-regulation of diverse cellular activities [ 29 ], whereby many genes and/or proteins clusters are strategically up-regulated or down-regulated to meet the challenges associated with aestivation. At present, we were unable to identify an appropriate reference protein, the expression of which would be unaffected throughout the three phases of aestivation, for Western blotting.…”

“…with reference to the total protein abundance, which has been reported previously for several other proteins from P . annectens during aestivation [ 24 , 29 – 34 ].…”

Rapidly evolving genes underlie Aedes aegypti mosquito reproductive resilience during drought

“…For example, when the river-dwelling African lungfish ( Protopterus annectens ) experiences food and water scarcity during drought, it burrows into the dried riverbed, forming a cocoon with secreted mucus. It can survive for years while remaining metabolically dormant but within a week of rainfall, it reawakens and resumes normal metabolism ( 1, 2 ). Among birds, species like the blackcap ( Sylvia atricapilla ) exhibit genetically-encoded seasonal migratory behaviors that rapidly evolve in the face of changing resource availability, resulting in new migratory routes and destinations ( 3–5 ).…”