Carbonic anhydrase XIV in skeletal muscle: subcellular localization and function from wild-type and knockout mice

Abstract

The expression of carbonic anhydrase (CA) XIV was investigated in mouse skeletal muscles. Sarcoplasmic reticulum (SR) and sarcolemmal (SL) membrane fractions were isolated from wild-type (WT) and CA XIV knockout (KO) mice. The CA XIV protein of 54 kDa was present in SR and SL membrane fractions as shown by Western blot analysis. CA activity measurements of WT and KO membrane fractions showed that CA XIV accounts for approximately 50% and 66% of the total CA activities determined in the SR and SL fractions, respectively. This indicates the presence of at least one other membrane-associated CA isoform in these membranes, e.g., CA IV, CA IX, or CA XII. Muscle fibers of the extensor digitorum longus (EDL) muscle were immunostained with anti-CA XIV/FITC and anti-sarco(endo)plasmic reticulum Ca(2+)-ATPase 1/TRITC, with anti-CA XIV/FITC and anti-ryanodine receptor/TRITC, or with anti-CA XIV/FITC and anti-monocarboxylate transporter-4/TRITC. CA XIV was expressed in the plasma membrane and in the longitudinal SR but not in the terminal SR. Isometric contraction measurements of single twitches and tetani and a fatigue protocol applied to fiber bundles of the fast-twitch EDL and of the slow-twitch soleus muscle from WT and KO mice showed that the lack of SR membrane-associated CA XIV did not affect maximum force, rise and relaxation times, and fatigue behavior. Thus, it is concluded that a reduction of the total SR CA activity by approximately 50% in CA XIV KO mice does not lead to an impairment of SR function.

T Tubules and Surface Membranes Provide Equally Effective Pathways of Carbonic Anhydrase-Facilitated Lactic Acid Transport in Skeletal Muscle

“…1b (lower lane) shows punctate staining, which, as also seen in the merged image, is completely interrupted wherever triads are located. This is most easily explained by a staining of the longitudinal or light SR but not of the terminal cisternae and is compatible with the previous observation of complete colocalization of CA XIV and SERCA staining [7]. Another clear-cut implication of these pictures is the complete absence of CA XIV from T tubules.…”

“…While the results presented in Figs. 1a and b largely confirm recent observations [6], [7], the following results of Fig. 1c lead to an even clearer interpretation of these data.…”

“…For that reason, single mouse EDL fibers were isolated, permeabilized with Triton X-100 and stained with primary antibodies raised against mouse CA IV, CA IX or CA XIV and fluorescently labelled secondary antibodies. We have shown previously that the antibodies against CA IV, CA IX and CA XIV are highly specific and that immunocytochemical staining is absent in muscle fibers of mice in which the CA isozyme considered had been knocked out [6], [7]. In addition, omission of the primary antibodies confirmed specificity of the CA as well as of the MCT4 and ryanodine receptor (RyR) staining.…”

The Most Recently Discovered Carbonic Anhydrase, CA XV, Is Expressed in the Thick Ascending Limb of Henle and in the Collecting Ducts of Mouse Kidney

“…The activity of murine CA IV is only 10-20% of that of human CA IV [26], which has raised the question of whether the two isozymes, CA XIV and XV, could compensate for this loss of activity in the rodent kidney. Unlike CA XV, which has become a pseudogene in humans and chimpanzees, CA XIV is present in several human tissues, including the heart, liver, skeletal muscle, and brain [29], [37], [38], [39], but the human enzyme has significantly lower activity than the mouse enzyme [40]. Although CA XIV has been partially localized to the same nephron segments where CA IV is also produced [5], [10], CA XIV showed no overlapping with CA XV in its distribution.…”

Carbonic Anhydrase XIV: Structure, Functions, and Potential Medical Applications

“…Indeed, abundant expression was found in the brain, heart, skeletal muscle, and liver. At a lower level, the expression of hCA XIV was also demonstrated in eyes, colon, small intestine, urinary bladder, bone osteoclasts, and spinal cord (3,(7)(8)(9)(10)(11)(12)(13)(14)(15) (Table 12.1).…”