Potassium [K+]-deficient diets have contributed to the global epidemic of hypertension and chronic kidney disease(CKD).Giventhelowcostandeaseofincreasingdietary[K+],moreresearchisneededtounderstand how[K+]imbalanceleadstothesediseases.Thekidneyshandle90%of[K+],andthedistalconvolutedtubule (DCT) acts as a [K+] sensor via the WNK-SPAK (With-No-Lysine/Ste20/SPS-1-related proline-alanine-rich protein kinase) pathway. Gain-of-function mutations to this pathway lead to severe hypertension and hyperkalemiabyactivationofthethiazide-sensitivesodium/chlorideco-transporter(NCC).Curiously,dietary[K+] depletion or loading causes the WNK-SPAK kinases to assemble into large DCT-specific cytoplasmic puncta, that are not seen in mice on normokalemic diets. For years, the structure and function of these condensates, whichwecall?WNKbodies?,remainedamystery.Dr.Boyd-Shiwarski?sinitialworkhasidentifiedthattheseDCT- specificpunctaare(i)dependentupontheexpressionofkidneyspecific(KS)-WNK1(ii)potassium-sensitive;?(iii) membrane-less;?(iv)notassociatedwithconventionalorganelles;?and(v)associatedwithWNK-SPAKproteins. Basedonthesefindings,wehypothesizethatWNKbodiesaremembrane-lessmicrodomainsthatsequesterthe WNK-SPAKpathwaytomodulateWNKsignalingduringpotassiumimbalance.Thishypothesiswillbetestedin twoaimsthatevaluatethephysiologicalsignificanceandbiologicalbasisofWNKbodyformation.Thisproposal?s physiology-based aim will provide Dr. Boyd-Shiwarski with the opportunity to work with animal models and (i) implement ex vivo microscopy techniques, (ii) quantify changes in urine, serum, and blood pressure, and (iii) develop transgenic mouse models. Whereas, the biology-based portion of this proposal will include implementation of (i) molecular cloning, (ii) protein biochemistry, and (iii) mass spectrometry and RNA Seq. Theseskillswillbereinforcedbyateamofmentors,advisors,collaborators,andcoreresourcesavailableatthe University of Pittsburgh. The primary mentor, Dr. Arohan Subramanya, is an established NIH R01-funded physician-scientist with 13 years of experience in WNK signaling biology and prior experience mentoring over 20 trainees. The co-mentor, Dr. Tom Kleyman, is an internationally recognized physician scientist who directs thePittsburghCenterforKidneyResearch,andhasmentoredninecareerdevelopmentawardeesandfiveR01 recipients within the last 10 years. In addition, an advisory committee of accomplished investigators with expertise in hypertension, WNK-SPAK signaling, and renal tubular transport will monitor Dr. Boyd-Shiwarski?s progressthroughbiannualmeetings.Dr.Boyd-Shiwarskiwillusethisproposaltoaccomplishhershort-termgoal ofscientificindependenceandherlong-termgoalofbecomingatenuretrackphysician-scientistwithexpertise inpotassiumhomeostasis,hypertension,andCKD.TheresultsfromthisproposalwillformthebasisforanR01 studyingthetranslationalroleofKS-WNK1-dependentWNKbodiesinhumanmodelsofnephropathy.

Public Health Relevance

98% of the US population does not meet the daily recommendation for dietary potassium intake, and this inadequatepotassiumintakeisdirectlylinkedtothedevelopmentofhypertensionandchronickidneydisease. The kidneys handle 90% of the serum potassium, however, the mechanism by which the kidneys sense and respond to potassium imbalance are poorly understood. This proposed study will advance our basic understandingofthesignalingpathwaysinthekidneythatrespondtopotassiumimbalance,andthusregulate bloodpressureandpotassiumhomeostasis.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08DK118211-02
Application #
9735226
Study Section
Kidney, Urologic and Hematologic Diseases D Subcommittee (DDK)
Program Officer
Rankin, Tracy L
Project Start
2018-07-01
Project End
2023-04-30
Budget Start
2019-05-01
Budget End
2020-04-30
Support Year
2
Fiscal Year
2019
Total Cost
Indirect Cost
Name
University of Pittsburgh
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213